Crystalline salt forms of mesembrine

ABSTRACT

The present invention relates to novel crystalline salt forms of mesembrine, also known as 3a-(3,4-dimethoxyphenyl)-octahydro-1-methy-6H-indol-6-one. Mesembrine has the chemical formula C17H23NO3. The invention further relates to the preparation of a novel crystalline salt of mesembrine and to the use of the mesembrine salt as a medicament. In one embodiment the novel crystalline salt form of mesembrine is mesembrine besylate salt.

The present application is related to, and claims the benefit ofGB2204778.1, filed on 1 Apr. 2022 (1 Apr. 2022), the contents of whichare hereby incorporated by reference in their entirety.

FIELD OF THE INVENTION

The present invention relates to novel crystalline salt forms ofmesembrine, also known as3a-(3,4-dimethoxyphenyl)-octahydro-1-methy-6H-indol-6-one. Mesembrinehas the chemical formula C₁₇H₂₃NO₃.

The invention further relates to the preparation of a novel crystallinesalt of mesembrine and to the use of the mesembrine salt as amedicament. In one embodiment the novel crystalline salt form ofmesembrine is mesembrine besylate salt.

BACKGROUND TO THE INVENTION

Mesembrine is an alkaloid which naturally occurs in the Sceletiumtortuosum species of plants indigenous to South Africa. The genusSceletium, classified under the Aizoaceae family, is indigenous to theWestern, Eastern and Northern Cape province of South Africa. In additionto mesembrine other alkaloids are found in extracts of Sceletiumtortuosum including mesembrenol, Δ⁷mesembrenone, mesembranol,mesembrenone, and epimesembranol.

Extracts of S. tortuosum have a long history of use in traditionalmedicine by the San and Khoikhoi people in South Africa where it wasused as a masticatory and a medicine to quench their thirst, fightfatigue and for healing, social, and spiritual purposes.

More recently studies have revealed that extracts of the plant havenumerous biological properties and extracts of S. tortuosum may beuseful in the treatment of anxiety and depression, psychological andpsychiatric disorders, improving mood, promoting relaxation andhappiness.

An in vivo study in rats demonstrated a positive effect of an extract ofS. tortuosum on restraint-induced anxiety (Smith, 2011), and a smallseries of case reports described preliminary evidence for antidepressantand anxiolytic activity in patients suffering from major depression whowere treated with tablets comprising a standardized extract of milled S.tortuosum raw material (Gericke, 2001). A dietary supplement comprisingsuch material is available as Zembrin®.

The mechanisms of action on the central nervous system (CNS) of Zembrin®were identified as the ability to cause blockade of the serotonin (5-HT)transporter and enable selective inhibition of the phosphodiesterase-4(PDE4) enzyme (Harvey et al, 2011).

The various alkaloids which occur in S. tortuosum have also been studiedin particular the three main alkaloids, mesembrenol, mesembrenone, andmesembrine. All three have been shown to be potently active in a 5-HTtransporter binding assay and against PDE4B activity, (Harvey et al.,2011).

Mesembrenone was described as having a dual activity on 5-HT uptake andPDE4 inhibition as the difference IC50 concentrations on the two assayswas x17, whereas it was x258 for mesembrenol and x5500 mesembrine.However, mesembrine had a greater selectivity for the 5-HT transporterover PDE4B.

The structure of mesembrine was described by Popelak et al., 1960 andthe configuration by P. W. Jeffs et al.,1969. Mesembrine occursnaturally as the (−)-isomer as (−)-mesembrine.

Mesembrine can be isolated from extracts of S. tortuosum or can besynthesized chemically using the method described by Wang et al., 2016.

Mesembrine has a solubility of 10 mg/ml in chloroform and ethanol buthas a low solubility in water meaning that it's use as in thepreparation of a pharmaceutical is limited as such formation of a salineform of the compound is desirable.

In 1957 the mesembrine base was successfully crystallized to itshydrochloride salt (Bodendorf and Krieger, 1957), however the HCl saltof mesembrine is poorly soluble at higher concentrations meaning onlysmall doses of mesembrine can be prepared using the HCl salt.

An object of the present invention is the preparation of a salt form ofmesembrine with superior properties to those presently available.

It has now been found that the monobenzenesulfonate (also referred to asbesylate) salified form of mesembrine exhibits advantageous propertieswhich render it particularly suitable for use as active principle in amedicament.

Specifically, the applicant has demonstrated, that the besylate form ofmesembrine unexpectedly has superior properties in comparison to othersalt forms. In particular the mesembrine besylate salt of the inventionhas improved solubility and stability, which are further improved withrespect to the hydrochloride or fumarate form of this same compound.

The advantages related to the besylate salt form of mesembrine incomparison to the free base form or to other saline forms, such as thehydrochloride and the fumarate, are described with reference tophysicochemical analysis and characterization.

BRIEF SUMMARY OF THE DISCLOSURE

In accordance with a first aspect of the present invention there isprovided a mesembrine salt, wherein the salt is taken from the groupconsisting of mesembrine besylate; mesembrine phosphate; mesembrinetartrate; mesembrine fumarate and mesembrine succinate.

Preferably the mesembrine salt is a mesembrine besylate salt.

More preferably the mesembrine salt is in a solid form. More preferablythe salt is in a crystalline form.

When the mesembrine salt of the invention in mesembrine besylate thesalt is preferably characterized by an XRPD pattern according to FIG. 18comprising peaks at about the positions as described in Table 3.10. Sucha salt is defined as mesembrine besylate salt Pattern 1.

In embodiments, the crystalline form is characterized by peaks in anXPRD pattern at 11.1±0.2, 12.7±0.2, 16.6±0.2, 23.8±0.2, and 24.6±0.2°2θ. In embodiments, the crystalline form is further characterized by atleast one peak selected from 9.2±0.2, 11.0±0.2, 13.5±0.2, 19.5±0.2,20.7±0.2, and 21.2±0.2°2θ. In embodiments, the variance at any of thesepeaks is ±0.1°2θ. For example, in embodiments, the crystalline form ischaracterized by peaks in an XPRD pattern at 11.1±0.1, 12.7±0.1,16.6±0.1, 23.8±0.1, and 24.6±0.1°2θ. In embodiments, the crystallineform is further characterized by at least one peak selected from9.2±0.1, 11.0±0.1, 13.5±0.1, 19.5±0.1, 20.7±0.1, and 21.2±0.1°2θ.

In embodiments, the crystalline form is characterized by peaks in a XRPDpattern at 9.2±0.2, 11.0±0.2, 11.1±0.2, 12.3±0.2, 12.7±0.2, 13.5±0.2,15.4±0.2, 16.6±0.2, 18.5±0.2, 19.5±0.2, 19.8±0.2, 20.2±0.2, 20.7±0.2,21.2±0.2, 21.6±0.2, 22.4±0.2, 22.9±0.2, 23.2±0.2, 23.8±0.2, 24.1±0.2,24.6±0.2, 25.6±0.2, 26.2±0.2, 27.9±0.2, 28.3±0.2, 28.6±0.2, 29.3±0.2,31.1±0.2, 32.4±0.2, 33.0±0.2, and 33.9±0.2°2θ. In embodiments, thevariance at any of these peaks is ±0.1°2θ.

In embodiments, the crystalline form is characterized by any one, two,three, four, five, six, seven, eight, nine, ten, eleven, or more, XRPDpeaks listed in Table 1.1.

TABLE 1.1 XPRD Peaks for the Mesmebrine Besylate Salt: Pattern 1 Peak #Pos. [° 2θ] 1 9.2 2 11.0 3 11.1 4 12.3 5 12.7 6 13.5 7 15.4 8 16.6 918.5 10 19.5 11 19.8 12 20.2 13 20.7 14 21.2 15 21.6 16 22.4 17 22.9 1823.2 19 23.8 20 24.1 21 24.6 22 25.6 23 26.2 24 27.9 25 28.3 26 28.6 2729.3 28 31.1 29 32.4 30 33.0 31 33.9

When the mesembrine salt of the invention in mesembrine besylate thesalt is alternatively characterized by an XRPD pattern according to FIG.20 comprising peaks at about the positions as described in Table 3.13.Such a salt is defined as mesembrine besylate salt Pattern 2.

In embodiments, the crystalline form is characterized by peaks in anXPRD pattern at 11.0±0.2, 13.4±0.2, 15.1±0.2, 18.6±0.2, or 23.7±0.2°2θ.In embodiments, the crystalline form is further characterized by atleast one peak selected from 15.6±0.2, 16.1±0.2, 18.2±0.2, 21.3±0.2, or25.1±0.2°2θ. In embodiments, the variance at any of these peaks is±0.1°2θ. For example, in embodiments, the crystalline form ischaracterized by peaks in an XPRD pattern at 11.0±0.1, 13.4±0.1,15.1±0.1, 18.6±0.1, or 23.7±0.1°2θ. In embodiments, the crystalline formis further characterized by at least one peak selected from 15.6±0.1,16.1±0.1, 18.2±0.1, 21.3±0.1, or 25.1±0.1°2θ.

In embodiments, the crystalline form is characterized by peaks in a XRPDpattern at 3.2±0.2, 7.4±0.2, 9.3±0.2, 11.0±0.2, 11.6±0.2, 12.1±0.2,12.6±0.2, 13.4±0.2, 14.5±0.2, 15.1±0.2, 15.6±0.2, 16.1±0.2, 16.8±0.2,17.9±0.2, 18.2±0.2, 18.6±0.2, 18.8±0.2, 19.5±0.2, 19.8±0.2, 20.7±0.2,21.3±0.2, 21.8±0.2, 22.4±0.2, 22.6±0.2, 22.7±0.2, 23.3±0.2, 23.7±0.2,24.1±0.2, 24.3±0.2, 24.7±0.2, 25.1±0.2, 25.7±0.2, 26.3±0.2, 26.6±0.2,27.0±0.2, 27.5±0.2, 28.0±0.2, 28.5±0.2, 29.0±0.2, 29.3±0.2, 29.5±0.2,30.0±0.2, 30.7±0.2, 31.7±0.2, 32.1±0.2, 32.5±0.2, 33.1±0.2, 33.5±0.2,and 34.4±0.2°2θ. In embodiments, the variance at any of these peaks is±0.1°2θ.

In embodiments, the crystalline form is characterized by any one, two,three, four, five, six, seven, eight, nine, ten, eleven, or more, XRPDpeaks listed in Table 1.2.

TABLE 1.2 XPRD Peaks for the Mesmebrine Besylate Salt: Pattern 2 Peak #Pos. [° 2θ] 1 3.2 2 7.4 3 9.3 4 11.0 5 11.6 6 12.1 7 12.6 8 13.4 9 14.510 15.1 11 15.6 12 16.1 13 16.8 14 17.9 15 18.2 16 18.6 17 18.8 18 19.519 19.8 20 20.7 21 21.3 22 21.8 23 22.4 24 22.6 25 22.7 26 23.3 27 23.728 24.1 29 24.3 30 24.7 31 25.1 32 25.7 33 26.3 34 26.6 35 27.0 36 27.537 28.0 38 28.5 39 29.0 40 29.3 41 29.5 42 30.0 43 30.7 44 31.7 45 32.146 32.5 47 33.1 48 33.5 49 34.4

In accordance with a second aspect of the present invention there isprovided a process for the preparation of a mesembrine salt comprisingthe steps of:

-   -   a) Dissolving mesembrine in a solvent    -   b) Addition of the appropriate counterion to the mesembrine        solution under temperature cycling conditions; and    -   c) Isolation of solids comprising the mesembrine salt.

In one embodiment the counterion of step b) is a weak acid with a pKa ofgreater than 0.5. Preferably the counterion of step b) isbenzenesulfonic acid.

In accordance with a third aspect of the present invention there isprovided a pharmaceutical preparation comprising a mesembrine salt,wherein the salt is taken from the group consisting of mesembrinebesylate; mesembrine phosphate; mesembrine tartrate; mesembrine fumarateand mesembrine succinate.

Preferably the salt is mesembrine besylate.

In accordance with a fourth aspect of the present invention there isprovided a mesembrine salt for use in the treatment of a disease,wherein the salt is taken from the group consisting of mesembrinebesylate; mesembrine phosphate; mesembrine tartrate; mesembrine fumarateand mesembrine succinate.

Preferably the mesembrine salt is mesembrine besylate.

The mesembrine salt of the invention may be used in the treatment ofimpulse control in a human or animal subject. In particular thetreatment may be related to the field of sexual dysfunction such asdelaying ejaculation, delaying orgasm and/or preventing prematureejaculation during sexual activity in human males. In addition themesembrine salt of the invention may be used to delay ejaculatorylatency in a male human.

The compositions provided herein contain therapeutically effectiveamounts of one or more of the compounds provided herein that are usefulin the prevention, treatment, or amelioration of one or more of thesymptoms of diseases or disorders described herein and a vehicle.Vehicles suitable for administration of the compounds provided hereininclude any such carriers known to those skilled in the art to besuitable for the particular mode of administration.

In addition, the compounds may be formulated as the sole activeingredient in the composition or may be combined with other activeingredients.

The compositions contain one or more compounds provided herein. Thecompounds are, in some embodiments, formulated into suitablepreparations such as solutions, suspensions, tablets, dispersibletablets, pills, capsules, powders, sustained release formulations orelixirs, for oral administration or in sterile solutions or suspensionsfor parenteral administration, as well as topical administration,transdermal administration, nasal inhalation, and oral inhalation vianebulizers, pressurized metered dose inhalers and dry powder inhalers.In some embodiments, the compounds described above are formulated intocompositions using techniques and procedures well known in the art (see,e.g., Ansel, Introduction to Pharmaceutical Dosage Forms, SeventhEdition (1999)).

In the compositions, effective concentrations of one or more compoundsor derivatives thereof is (are) mixed with a suitable vehicle. Thecompounds may be derivatized as the corresponding salts, esters, enolethers or esters, acetals, ketals, orthoesters, hemiacetals, hemiketals,acids, bases, solvates, ion-pairs, hydrates or prodrugs prior toformulation, as described above. The concentrations of the compounds inthe compositions are effective for delivery of an amount, uponadministration that treats, leads to prevention, or amelioration of oneor more of the symptoms of diseases or disorders described herein. Insome embodiments, the compositions are formulated for single dosageadministration. To formulate a composition, the weight fraction of acompound is dissolved, suspended, dispersed or otherwise mixed in aselected vehicle at an effective concentration such that the treatedcondition is relieved, prevented, or one or more symptoms areameliorated.

The active compound is included in the vehicle in an amount sufficientto exert a therapeutically useful effect in the absence of undesirableside effects on the patient treated. The therapeutically effectiveconcentration may be predicted empirically by testing the compounds inin vitro and in vivo systems well known to those of skill in the art andthen extrapolated therefrom for dosages for humans. Human doses are thentypically fine-tuned in clinical trials and titrated to response.

The concentration of active compound in the composition will depend onabsorption, inactivation and excretion rates of the active compound, thephysicochemical characteristics of the compound, the dosage schedule,and amount administered as well as other factors known to those of skillin the art. For example, the amount that is delivered is sufficient toameliorate one or more of the symptoms of diseases or disorders asdescribed herein.

In some embodiments, a therapeutically effective dosage should produce aserum concentration of active ingredient of from about 0.001 ng/ml toabout 1.0 ng/ml, 2-10 ng/ml, 11 to 50 ng/ml, 51-200 ng/ml, or about 200to 1000 ng/ml. The compositions, in other embodiments, should provide adosage of from about 0.0001 mg to about 70 mg of compound per kilogramof body weight per day. Dosage unit forms are prepared to provide fromabout 0.01 mg, 0.1 mg or 1 mg to about 500 mg, or about 1000 mg, and insome embodiments from about 10 mg to about 500 mg of the activeingredient or a combination of essential ingredients per dosage unitform.

The active ingredient may be administered at once or may be divided intoa number of smaller doses to be administered at intervals of time. It isunderstood that the precise dosage and duration of treatment is afunction of the disease being treated and may be determined empiricallyusing known testing protocols or by extrapolation from in vivo or invitro test data or subsequent clinical testing. It is to be noted thatconcentrations and dosage values may also vary with the severity of thecondition to be alleviated. It is to be further understood that for anyparticular subject, specific dosage regimens should be adjusted overtime according to the individual need and the professional judgment ofthe person administering or supervising the administration of thecompositions and that the concentration ranges set forth herein areexemplary only and are not intended to limit the scope or practice ofthe claimed compositions.

In instances in which the compounds exhibit insufficient solubility,methods for solubilizing compounds may be used such as use of liposomes,prodrugs, complexation/chelation, nanoparticles, or emulsions ortertiary templating. Such methods are known to those of skill in thisart, and include, but are not limited to, using co-solvents, such asdimethylsulfoxide (DMSO), using surfactants or surface modifiers, suchas TWEEN®, complexing agents such as cyclodextrin or dissolution byenhanced ionization (i.e., dissolving in aqueous sodium bicarbonate).Derivatives of the compounds, such as prodrugs of the compounds may alsobe used in formulating effective compositions.

Upon mixing or addition of the compound(s), the resulting mixture may bea solution, suspension, emulsion or the like. The form of the resultingmixture depends upon a number of factors, including the intended mode ofadministration and the solubility of the compound in the selectedvehicle. The effective concentration is sufficient for ameliorating thesymptoms of the disease, disorder or condition treated and may beempirically determined.

The compositions are provided for administration to humans and animalsin indication appropriate dosage forms, such as dry powder inhalers(DPIs), pressurized metered dose inhalers (pMDIs), nebulizers, tablets,capsules, pills, sublingual tapes/bioerodible strips, tablets orcapsules, powders, granules, lozenges, lotions, salves, suppositories,fast melts, transdermal patches or other transdermal applicationdevices/preparations, sterile parenteral solutions or suspensions, andoral solutions or suspensions, and oil-water emulsions containingsuitable quantities of the compounds or derivatives thereof. Thetherapeutically active compounds and derivatives thereof are, in someembodiments, formulated and administered in unit-dosage forms ormultiple-dosage forms. Unit-dose forms as used herein refer tophysically discrete units suitable for human and animal subjects andpackaged individually as is known in the art. Each unit-dose contains apredetermined quantity of the therapeutically active compound sufficientto produce the desired therapeutic effect, in association with therequired vehicle. Examples of unit-dose forms include ampoules andsyringes and individually packaged tablets or capsules. Unit-dose formsmay be administered in fractions or multiples thereof. A multiple-doseform is a plurality of identical unit-dosage forms packaged in a singlecontainer to be administered in segregated unit-dose form. Examples ofmultiple-dose forms include vials, bottles of tablets or capsules orbottles of pints or gallons. Hence, multiple dose form is a multiple ofunit-doses which are not segregated in packaging.

Liquid compositions can, for example, be prepared by dissolving,dispersing, or otherwise mixing an active compound as defined above andoptional adjuvants in a vehicle, such as, for example, water, saline,aqueous dextrose, glycerol, glycols, ethanol, and the like, to therebyform a solution or suspension, colloidal dispersion, emulsion orliposomal formulation. If desired, the composition to be administeredmay also contain minor amounts of nontoxic auxiliary substances such aswetting agents, emulsifying agents, solubilizing agents, pH bufferingagents and the like, for example, acetate, sodium citrate, cyclodextrinderivatives, sorbitan monolaurate, triethanolamine sodium acetate,triethanolamine oleate, and other such agents.

Actual methods of preparing such dosage forms are known, or will beapparent, to those skilled in this art; for example, see Remington'sPharmaceutical Sciences, Mack Publishing Company, Easton, Pa., 15thEdition, 1975 or later editions thereof.

Dosage forms or compositions containing active ingredient in the rangeof 0.005% to 100% with the balance made up from vehicle or carrier maybe prepared. Methods for preparation of these compositions are known tothose skilled in the art. The contemplated compositions may contain0.001%-100% active ingredient, in one embodiment 0.1-95%, in anotherembodiment 0.4-10%.

In certain embodiments, the compositions are lactose-free compositionscontaining excipients that are well known in the art and are listed, forexample, in the U.S. Pharmacopeia (USP) 25-NF20 (2002). In general,lactose-free compositions contain active ingredients, a binder/filler,and a lubricant in compatible amounts. Particular lactose-free dosageforms contain active ingredients, microcrystalline cellulose,pre-gelatinized starch, and magnesium stearate.

Further provided are anhydrous compositions and dosage forms includingactive ingredients, since water can facilitate the degradation of somecompounds. For example, the addition of water (e.g., 5%) is widelyaccepted as a means of simulating long-term storage in order todetermine characteristics such as shelf-life or the stability offormulations over time. See, e.g., Jens T. Carstensen, Drug Stability:Principles & Practice, 2d. Ed., Marcel Dekker, NY, N.Y., 1995, pp.379-80. In effect, water and heat accelerate the decomposition of somecompounds. Thus, the effect of water on a formulation can be of greatsignificance since moisture and/or humidity are commonly encounteredduring manufacture, handling, packaging, storage, shipment, and use offormulations.

Anhydrous compositions and dosage forms provided herein can be preparedusing anhydrous or low moisture containing ingredients and low moistureor low humidity conditions.

An anhydrous composition should be prepared and stored such that itsanhydrous nature is maintained. Accordingly, anhydrous compositions aregenerally packaged using materials known to prevent exposure to watersuch that they can be included in suitable formulary kits. Examples ofsuitable packaging include, but are not limited to, hermetically sealedfoils, plastics, unit dose containers (e.g., vials), blister packs, andstrip packs.

Oral dosage forms are either solid, gel or liquid. The solid dosageforms are tablets, capsules, granules, and bulk powders. Types of oraltablets include compressed, chewable lozenges and tablets which may beenteric-coated, sugar-coated or film-coated. Capsules may be hard orsoft gelatin capsules, while granules and powders may be provided innon-effervescent or effervescent form with the combination of otheringredients known to those skilled in the art.

In certain embodiments, the formulations are solid dosage forms such asfor example, capsules or tablets. The tablets, pills, capsules, trochesand the like can contain one or more of the following ingredients, orcompounds of a similar nature: a binder; a lubricant; a diluent; aglidant; a disintegrating agent; a coloring agent; a sweetening agent; aflavoring agent; a wetting agent; an enteric coating; a film coatingagent and modified release agent. Examples of binders includemicrocrystalline cellulose, methyl paraben, polyalkyleneoxides, gumtragacanth, glucose solution, acacia mucilage, gelatin solution,molasses, polyvinylpyrrolidine, povidone, crospovidones, sucrose andstarch and starch derivatives. Lubricants include talc, starch,magnesium/calcium stearate, lycopodium and stearic acid. Diluentsinclude, for example, lactose, sucrose, trehalose, lysine, leucine,lecithin, starch, kaolin, salt, mannitol and dicalcium phosphate.Glidants include, but are not limited to, colloidal silicon dioxide.Disintegrating agents include crosscarmellose sodium, sodium starchglycolate, alginic acid, corn starch, potato starch, bentonite,methylcellulose, agar and carboxymethylcellulose. Coloring agentsinclude, for example, any of the approved certified water-soluble FD andC dyes, mixtures thereof; and water insoluble FD and C dyes suspended onalumina hydrate and advanced coloring or anti-forgery color/opalescentadditives known to those skilled in the art. Sweetening agents includesucrose, lactose, mannitol and artificial sweetening agents such assaccharin, and any number of spray dried flavors. Flavoring agentsinclude natural flavors extracted from plants such as fruits andsynthetic blends of compounds which produce a pleasant sensation or maskunpleasant taste, such as, but not limited to peppermint and methylsalicylate. Wetting agents include propylene glycol monostearate,sorbitan monooleate, diethylene glycol monolaurate and polyoxyethylenelauryl ether. Enteric coatings include fatty acids, fats, waxes,shellac, ammoniated shellac and cellulose acetate phthalates. Filmcoatings include hydroxyethylcellulose, sodium carboxymethylcellulose,polyethylene glycol 4000 and cellulose acetate phthalate. Modifiedrelease agents include polymers such as the Eudragit° series andcellulose esters.

The compound, or derivative thereof, can be provided in a compositionthat protects it from the acidic environment of the stomach. Forexample, the composition can be formulated in an enteric coating thatmaintains its integrity in the stomach and releases the active compoundin the intestine. The composition may also be formulated in combinationwith an antacid or other such ingredient.

When the dosage unit form is a capsule, it can contain, in addition tomaterial of the above type, a liquid carrier such as a fatty oil. Inaddition, dosage unit forms can contain various other materials whichmodify the physical form of the dosage unit, for example, coatings ofsugar and other enteric agents. The compounds can also be administeredas a component of an elixir, suspension, syrup, wafer, sprinkle, chewinggum or the like. A syrup may contain, in addition to the activecompounds, sucrose as a sweetening agent and certain preservatives, dyesand colorings and flavors.

The active materials can also be mixed with other active materials whichdo not impair the desired action, or with materials that supplement thedesired action. The active ingredient is a compound or derivativethereof as described herein. Higher concentrations, up to about 98% byweight of the active ingredient may be included.

In all embodiments, tablets and capsules formulations may be coated asknown by those of skill in the art in order to modify or sustaindissolution of the active ingredient. Thus, for example, they may becoated with a conventional enterically digestible coating, such asphenylsalicylate, waxes and cellulose acetate phthalate.

Liquid oral dosage forms include aqueous solutions, emulsions,suspensions, solutions and/or suspensions reconstituted fromnon-effervescent granules and effervescent preparations reconstitutedfrom effervescent granules. Aqueous solutions include, for example,elixirs and syrups. Emulsions are either oil-in-water or water-in-oil.

Elixirs are clear, sweetened, hydroalcoholic preparations. Vehicles usedin elixirs include solvents. Syrups are concentrated aqueous solutionsof a sugar, for example, sucrose, and may contain a preservative. Anemulsion is a two-phase system in which one liquid is dispersed in theform of small globules throughout another liquid. Carriers used inemulsions are non-aqueous liquids, emulsifying agents and preservatives.Suspensions use suspending agents and preservatives. Acceptablesubstances used in non-effervescent granules, to be reconstituted into aliquid oral dosage form, include diluents, sweeteners and wettingagents. Acceptable substances used in effervescent granules, to bereconstituted into a liquid oral dosage form, include organic acids anda source of carbon dioxide. Coloring and flavoring agents are used inall of the above dosage forms.

Solvents include glycerin, sorbitol, ethyl alcohol and syrup. Examplesof preservatives include glycerin, methyl and propylparaben, benzoicacid, sodium benzoate and alcohol. Examples of non-aqueous liquidsutilized in emulsions include mineral oil and cottonseed oil. Examplesof emulsifying agents include gelatin, acacia, tragacanth, bentonite,and surfactants such as polyoxyethylene sorbitan monooleate. Suspendingagents include sodium carboxymethylcellulose, pectin, tragacanth, Veegumand acacia. Sweetening agents include sucrose, syrups, glycerin andartificial sweetening agents such as saccharin. Wetting agents includepropylene glycol monostearate, sorbitan monooleate, diethylene glycolmonolaurate and polyoxyethylene lauryl ether. Organic acids includecitric and tartaric acid. Sources of carbon dioxide include sodiumbicarbonate and sodium carbonate. Coloring agents include any of theapproved certified water-soluble FD and C dyes, and mixtures thereof.Flavoring agents include natural flavors extracted from plants suchfruits, and synthetic blends of compounds, which produce a pleasanttaste sensation.

For a solid dosage form, the solution or suspension, in for example,propylene carbonate, vegetable oils or triglycerides, is in someembodiments encapsulated in a gelatin capsule. Such solutions, and thepreparation and encapsulation thereof, are disclosed in U.S. Pat. Nos.4,328,245; 4,409,239; and 4,410,545. For a liquid dosage form, thesolution, e.g., for example, in a polyethylene glycol, may be dilutedwith a sufficient quantity of a liquid vehicle, e.g., water, to beeasily measured for administration.

Alternatively, liquid or semi-solid oral formulations may be prepared bydissolving or dispersing the active compound or salt in vegetable oils,glycols, triglycerides, propylene glycol esters (e.g., propylenecarbonate) and other such carriers, and encapsulating these solutions orsuspensions in hard or soft gelatin capsule shells. Other usefulformulations include those set forth in U.S. Pat. Nos. RE 28,819 and4,358,603. Briefly, such formulations include, but are not limited to,those containing a compound provided herein, a dialkylated mono- orpolyalkylene glycol, including, but not limited to, 1,2-dimethoxyethane,diglyme, triglyme, tetraglyme, polyethylene glycol-350-dimethyl ether,polyethylene glycol-550-dimethyl ether, polyethylene glycol-750-dimethylether wherein 350, 550 and 750 refer to the approximate averagemolecular weight of the polyethylene glycol, and one or moreantioxidants, such as butylated hydroxytoluene (BHT), butylatedhydroxyanisole (BHA), propyl gallate, vitamin E, hydroquinone,hydroxycoumarins, ethanolamine, lecithin, cephalin, ascorbic acid, malicacid, sorbitol, phosphoric acid, thiodipropionic acid and its esters,and dithiocarbamates.

Other formulations include, but are not limited to, aqueous alcoholicsolutions including an acetal. Alcohols used in these formulations areany water-miscible solvents having one or more hydroxyl groups,including, but not limited to, propylene glycol and ethanol. Acetalsinclude, but are not limited to, di(lower alkyl) acetals of lower alkylaldehydes such as acetaldehyde diethyl acetal.

Parenteral administration, in some embodiments characterized byinjection, either subcutaneously, intramuscularly or intravenously isalso contemplated herein. Injectables can be prepared in conventionalforms, either as liquid solutions or suspensions, solid forms suitablefor solution or suspension in liquid prior to injection, or asemulsions. The injectables, solutions and emulsions also contain one ormore excipients. Suitable excipients are, for example, water, saline,dextrose, glycerol or ethanol. In addition, if desired, the compositionsto be administered may also contain minor amounts of non-toxic auxiliarysubstances such as wetting or emulsifying agents, pH buffering agents,stabilizers, solubility enhancers, and other such agents, such as forexample, sodium acetate, sorbitan monolaurate, triethanolamine oleateand cyclodextrins.

Implantation of a slow-release or sustained-release system, such that aconstant level of dosage is maintained (see, e.g., U.S. Pat. No.3,710,795) is also contemplated herein. Briefly, a compound providedherein is dispersed in a solid inner matrix, e.g.,polymethylmethacrylate, polybutylmethacrylate, plasticized orunplasticized polyvinylchloride, plasticized nylon, plasticizedpolyethyleneterephthalate, natural rubber, polyisoprene,polyisobutylene, polybutadiene, polyethylene, ethylene-vinylacetatecopolymers, silicone rubbers, polydimethylsiloxanes, silicone carbonatecopolymers, hydrophilic polymers such as hydrogels of esters of acrylicand methacrylic acid, collagen, cross-linked polyvinylalcohol andcross-linked partially hydrolyzed polyvinyl acetate, that is surroundedby an outer polymeric membrane, e.g., polyethylene, polypropylene,ethylene/propylene copolymers, ethylene/ethyl acrylate copolymers,ethylene/vinylacetate copolymers, silicone rubbers, polydimethylsiloxanes, neoprene rubber, chlorinated polyethylene, polyvinylchloride,vinylchloride copolymers with vinyl acetate, vinylidene chloride,ethylene and propylene, ionomer polyethylene terephthalate, butyl rubberepichlorohydrin rubbers, ethylene/vinyl alcohol copolymer,ethylene/vinyl acetate/vinyl alcohol terpolymer, andethylene/vinyloxyethanol copolymer, that is insoluble in body fluids.The compound diffuses through the outer polymeric membrane in a releaserate controlling step. The percentage of active compound contained insuch parenteral compositions is highly dependent on the specific naturethereof, as well as the activity of the compound and the needs of thesubject.

Parenteral administration of the compositions includes intravenous,subcutaneous and intramuscular administrations. Preparations forparenteral administration include sterile solutions ready for injection,sterile dry soluble products, such as lyophilized powders, ready to becombined with a solvent just prior to use, including hypodermic tablets,sterile suspensions ready for injection, sterile dry insoluble productsready to be combined with a vehicle just prior to use and sterileemulsions. The solutions may be either aqueous or nonaqueous.

If administered intravenously, suitable carriers include physiologicalsaline or phosphate buffered saline (PBS), and solutions containingthickening and solubilizing agents, such as glucose, polyethyleneglycol, and polypropylene glycol and mixtures thereof.

Vehicles used in parenteral preparations include aqueous vehicles,nonaqueous vehicles, antimicrobial agents, isotonic agents, buffers,antioxidants, local anesthetics, suspending and dispersing agents,emulsifying agents, sequestering or chelating agents and othersubstances.

Examples of aqueous vehicles include Sodium Chloride Injection, RingersInjection, Isotonic Dextrose Injection, Sterile Water Injection,Dextrose and Lactated Ringers Injection. Nonaqueous parenteral vehiclesinclude fixed oils of vegetable origin, cottonseed oil, corn oil, sesameoil and peanut oil. Antimicrobial agents in bacteriostatic orfungistatic concentrations must be added to parenteral preparationspackaged in multiple-dose containers which include phenols or cresols,mercurials, benzyl alcohol, chlorobutanol, methyl and propylp-hydroxybenzoic acid esters, thimerosal, benzalkonium chloride andbenzethonium chloride. Isotonic agents include sodium chloride anddextrose. Buffers include phosphate and citrate. Antioxidants includesodium bisulfate. Local anesthetics include procaine hydrochloride.Suspending and dispersing agents include sodium carboxymethylcellulose,hydroxypropyl methylcellulose and polyvinylpyrrolidone. Emulsifyingagents include Polysorbate 80 (Tween® 80). A sequestering or chelatingagent of metal ions includes EDTA. Carriers also include ethyl alcohol,polyethylene glycol and propylene glycol for water miscible vehicles,and sodium hydroxide, hydrochloric acid, citric acid or lactic acid forpH adjustment.

The concentration of compound is adjusted so that an injection providesan effective amount to produce the desired pharmacological effect. Theexact dose depends on the age, weight, body surface area and conditionof the patient or animal as is known in the art.

The unit-dose parenteral preparations are packaged in an ampoule, a vialor a syringe with a needle. All preparations for parenteraladministration must be sterile, as is known and practiced in the art.

Illustratively, intravenous or intraarterial infusion of a sterileaqueous solution containing an active compound is an effective mode ofadministration. Another embodiment is a sterile aqueous or oily solutionor suspension containing an active material injected as necessary toproduce the desired pharmacological effect.

Injectables are designed for local and systemic administration. In someembodiments, a therapeutically effective dosage is formulated to containa concentration of at least about 0.01% w/w up to about 90% w/w or more,in certain embodiments more than 0.1% w/w of the active compound to thetreated tissue(s).

The compound may be suspended in micronized or other suitable form ormay be derivatized to produce a more soluble active product or toproduce a prodrug. The form of the resulting mixture depends upon anumber of factors, including the intended mode of administration and thesolubility of the compound in the selected carrier or vehicle. Theeffective concentration is sufficient for ameliorating the symptoms ofthe condition and may be empirically determined.

Active ingredients provided herein can be administered by controlledrelease means or by delivery devices that are well known to those ofordinary skill in the art. Examples include, but are not limited to,those described in U.S. Pat. Nos.: 3,845,770; 3,916,899; 3,536,809;3,598,123; 4,008,719; 5,674,533; 5,059,595; 5,591,767; 5,120,548;5,073,543; 5,639,476; 5,354,556; 5,639,480; 5,733,566; 5,739,108;5,891,474; 5,922,356; 5,972,891; 5,980,945; 5,993,855; 6,045,830;6,087,324; 6,113,943; 6,197,350; 6,248,363; 6,264,970; 6,267,981;6,376,461; 6,419,961; 6,589,548; 6,613,358; 6,699,500 and 6,740,634.Such dosage forms can be used to provide slow or controlled release ofone or more active ingredients using, for example, hydroxypropylmethylcellulose, other polymer matrices, gels, permeable membranes, osmoticsystems, multilayer coatings, microparticles, liposomes, microspheres,or a combination thereof to provide the desired release profile invarying proportions. Suitable controlled-release formulations known tothose of ordinary skill in the art, including those described herein,can be readily selected for use with the active ingredients providedherein.

All controlled-release products have a common goal of improving drugtherapy over that achieved by their non-controlled counterparts.Ideally, the use of an optimally designed controlled-release preparationin medical treatment is characterized by a minimum of drug substancebeing employed to cure or control the condition in a minimum amount oftime. Advantages of controlled-release formulations include extendedactivity of the drug, reduced dosage frequency, and increased patientcompliance. In addition, controlled-release formulations can be used toaffect the time of onset of action or other characteristics, such asblood levels of the drug, and can thus affect the occurrence of side(e.g., adverse) effects.

Most controlled-release formulations are designed to initially releasean amount of drug (active ingredient) that promptly produces the desiredtherapeutic effect, and gradually and continually release of otheramounts of drug to maintain this level of therapeutic or prophylacticeffect over an extended period of time. In order to maintain thisconstant level of drug in the body, the drug must be released from thedosage form at a rate that will replace the amount of drug beingmetabolized and excreted from the body. Controlled release of an activeingredient can be stimulated by various conditions including, but notlimited to, pH, temperature, enzymes, water, or other physiologicalconditions or compounds.

In certain embodiments, the agent may be administered using intravenousinfusion, an implantable osmotic pump, a transdermal patch, liposomes,or other modes of administration. In some embodiments, a pump may beused (see, Sefton, CRC Crit. Ref. Biomed. Eng. 14:201 (1987); Buchwaldet al., Surgery 88:507 (1980); Saudek et al., N Engl. J. Med. 321:574(1989)). In other embodiments, polymeric materials can be used. In otherembodiments, a controlled release system can be placed in proximity ofthe therapeutic target, i.e., thus requiring only a fraction of thesystemic dose (see, e.g., Goodson, Medical Applications of ControlledRelease, vol. 2, pp. 115-138 (1984)). In some embodiments, a controlledrelease device is introduced into a subject in proximity of the site ofinappropriate immune activation or a tumor. Other controlled releasesystems are discussed in the review by Langer (Science 249:1527-1533(1990)). The active ingredient can be dispersed in a solid inner matrix,e.g., polymethylmethacrylate, polybutylmethacrylate, plasticized orunplasticized polyvinylchloride, plasticized nylon, plasticizedpolyethyleneterephthalate, natural rubber, polyisoprene,polyisobutylene, polybutadiene, polyethylene, ethylene-vinylacetatecopolymers, silicone rubbers, polydimethylsiloxanes, silicone carbonatecopolymers, hydrophilic polymers such as hydrogels of esters of acrylicand methacrylic acid, collagen, cross-linked polyvinylalcohol andcross-linked partially hydrolyzed polyvinyl acetate, that is surroundedby an outer polymeric membrane, e.g., polyethylene, polypropylene,ethylene/propylene copolymers, ethylene/ethyl acrylate copolymers,ethylene/vinylacetate copolymers, silicone rubbers, polydimethylsiloxanes, neoprene rubber, chlorinated polyethylene, polyvinylchloride,vinylchloride copolymers with vinyl acetate, vinylidene chloride,ethylene and propylene, ionomer polyethylene terephthalate, butyl rubberepichlorohydrin rubbers, ethylene/vinyl alcohol copolymer,ethylene/vinyl acetate/vinyl alcohol terpolymer, andethylene/vinyloxyethanol copolymer, that is insoluble in body fluids.The active ingredient then diffuses through the outer polymeric membranein a release rate controlling step. The percentage of active ingredientcontained in such parenteral compositions is highly dependent on thespecific nature thereof, as well as the needs of the subject.

Of interest herein are also lyophilized powders, which can bereconstituted for administration as solutions, emulsions and othermixtures. They may also be reconstituted and formulated as solids orgels.

The sterile, lyophilized powder is prepared by dissolving a compoundprovided herein, or a derivative thereof, in a suitable solvent. Thesolvent may contain an excipient which improves the stability or otherpharmacological component of the powder or reconstituted solution,prepared from the powder. Excipients that may be used include, but arenot limited to, an antioxidant, a buffer and a bulking agent. In someembodiments, the excipient is selected from dextrose, sorbitol,fructose, corn syrup, xylitol, glycerin, glucose, sucrose and othersuitable agent. The solvent may contain a buffer, such as citrate,sodium or potassium phosphate or other such buffer known to those ofskill in the art at, at about neutral pH. Subsequent sterile filtrationof the solution followed by lyophilization under standard conditionsknown to those of skill in the art provides the desired formulation. Insome embodiments, the resulting solution will be apportioned into vialsfor lyophilization. Each vial will contain a single dosage or multipledosages of the compound. The lyophilized powder can be stored underappropriate conditions, such as at about 4° C. to room temperature.

Reconstitution of this lyophilized powder with water for injectionprovides a formulation for use in parenteral administration. Forreconstitution, the lyophilized powder is added to sterile water orother suitable carrier. The precise amount depends upon the selectedcompound. Such amount can be empirically determined.

Topical mixtures are prepared as described for the local and systemicadministration. The resulting mixture may be a solution, suspension,emulsions or the like and are formulated as creams, gels, ointments,emulsions, solutions, elixirs, lotions, suspensions, tinctures, pastes,foams, aerosols, irrigations, sprays, suppositories, bandages, dermalpatches or any other formulations suitable for topical administration.

The compounds or derivatives thereof may be formulated as aerosols fortopical application, such as by inhalation (see, e.g., U.S. Pat. Nos.4,044,126, 4,414,209, and 4,364,923, which describe aerosols fordelivery of a steroid useful for treatment of inflammatory diseases,particularly asthma). These formulations for administration to therespiratory tract can be in the form of an aerosol or solution for anebulizer, or as a microfine powder for insufflation, alone or incombination with an inert carrier such as lactose. In such a case, theparticles of the formulation will, in some embodiments, have mass mediangeometric diameters of less than 5 microns, in other embodiments lessthan 10 microns.

Oral inhalation formulations of the compounds or derivatives suitablefor inhalation include metered dose inhalers, dry powder inhalers andliquid preparations for administration from a nebulizer or metered doseliquid dispensing system. For both metered dose inhalers and dry powderinhalers, a crystalline form of the compounds or derivatives is thepreferred physical form of the drug to confer longer product stability.

In addition to particle size reduction methods known to those skilled inthe art, crystalline particles of the compounds or derivatives can begenerated using supercritical fluid processing which offers significantadvantages in the production of such particles for inhalation deliveryby producing respirable particles of the desired size in a single step.(e.g., International Publication No. WO2005/025506). A controlledparticle size for the microcrystals can be selected to ensure that asignificant fraction of the compounds or derivatives is deposited in thelung. In some embodiments, these particles have a mass medianaerodynamic diameter of about 0.1 to about 10 microns, in otherembodiments, about 1 to about 5 microns and still other embodiments,about 1.2 to about 3 microns.

Inert and non-flammable HFA propellants are selected from HFA 134a(1,1,1,2-tetrafluoroethane) and HFA 227e(1,1,1,2,3,3,3-heptafluoropropane) and provided either alone or as aratio to match the density of crystal particles of the compounds orderivatives. A ratio is also selected to ensure that the productsuspension avoids detrimental sedimentation or cream (which canprecipitate irreversible agglomeration) and instead promote a looselyflocculated system, which is easily dispersed when shaken. Looselyfluctuated systems are well regarded to provide optimal stability forpMDI canisters. As a result of the formulation's properties, theformulation contained no ethanol and no surfactants/stabilizing agents.

The compounds may be formulated for local or topical application, suchas for topical application to the skin and mucous membranes, such as inthe eye, in the form of gels, creams, and lotions and for application tothe eye or for intracisternal or intraspinal application. Topicaladministration is contemplated for transdermal delivery and also foradministration to the eyes or mucosa, or for inhalation therapies. Nasalsolutions of the active compound alone or in combination with otherexcipients can also be administered.

For nasal administration, the preparation may contain an esterifiedphosphonate compound dissolved or suspended in a liquid carrier, inparticular, an aqueous carrier, for aerosol application. The carrier maycontain solubilizing or suspending agents such as propylene glycol,surfactants, absorption enhancers such as lecithin or cyclodextrin, orpreservatives.

Solutions, particularly those intended for ophthalmic use, may beformulated as 0.01%-10% isotonic solutions, pH about 5-7.4, withappropriate salts.

Other routes of administration, such as transdermal patches, includingiontophoretic and electrophoretic devices, and rectal administration,are also contemplated herein.

Transdermal patches, including iontophoretic and electrophoreticdevices, are well known to those of skill in the art. For example, suchpatches are disclosed in U.S. Pat. Nos. 6,267,983, 6,261,595, 6,256,533,6,167,301, 6,024,975, 6,010715, 5,985,317, 5,983,134, 5,948,433 and5,860,957.

For example, dosage forms for rectal administration are rectalsuppositories, capsules and tablets for systemic effect. Rectalsuppositories are used herein mean solid bodies for insertion into therectum which melt or soften at body temperature releasing one or morepharmacologically or therapeutically active ingredients. Substancesutilized in rectal suppositories are bases or vehicles and agents toraise the melting point. Examples of bases include cocoa butter(theobroma oil), glycerin-gelatin, carbowax (polyoxyethylene glycol) andappropriate mixtures of mono-, di- and triglycerides of fatty acids.Combinations of the various bases may be used. Agents to raise themelting point of suppositories include spermaceti and wax. Rectalsuppositories may be prepared either by the compressed method or bymolding. The weight of a rectal suppository, in one embodiment, is about2 to 3 gm. Tablets and capsules for rectal administration aremanufactured using the same substance and by the same methods as forformulations for oral administration.

The compounds provided herein, or derivatives thereof, may also beformulated to be targeted to a particular tissue, receptor, or otherarea of the body of the subject to be treated. Many such targetingmethods are well known to those of skill in the art. All such targetingmethods are contemplated herein for use in the instant compositions. Fornon-limiting examples of targeting methods, see, e.g., U.S. Pat. Nos.6,316,652, 6,274,552, 6,271,359, 6,253,872, 6,139,865, 6,131,570,6,120,751, 6,071,495, 6,060,082, 6,048,736, 6,039,975, 6,004,534,5,985,307, 5,972,366, 5,900,252, 5,840,674, 5,759,542 and 5,709,874.

In some embodiments, liposomal suspensions, including tissue-targetedliposomes, such as tumor-targeted liposomes, may also be suitable ascarriers. These may be prepared according to methods known to thoseskilled in the art. For example, liposome formulations may be preparedas described in U.S. Pat. No. 4,522,811. Briefly, liposomes such asmultilamellar vesicles (MLV's) may be formed by drying down phosphatidylcholine and phosphatidyl serine (7:3 molar ratio) on the inside of aflask. A solution of a compound provided herein in phosphate bufferedsaline lacking divalent cations (PBS) is added and the flask shakenuntil the lipid film is dispersed. The resulting vesicles are washed toremove unencapsulated compound, pelleted by centrifugation, and thenresuspended in PBS.

The compounds or derivatives may be packaged as articles of manufacturecontaining packaging material, a compound or derivative thereof providedherein, which is effective for treatment, prevention or amelioration ofone or more symptoms of the diseases or disorders, supra, within thepackaging material, and a label that indicates that the compound orcomposition or derivative thereof, is used for the treatment, preventionor amelioration of one or more symptoms of the diseases or disorders,supra.

The articles of manufacture provided herein contain packaging materials.Packaging materials for use in packaging products are well known tothose of skill in the art. See, e.g., U.S. Pat. Nos. 5,323,907,5,052,558 and 5,033,252. Examples of packaging materials include, butare not limited to, blister packs, bottles, tubes, inhalers, pumps,bags, vials, containers, syringes, bottles, and any packaging materialsuitable for a selected formulation and intended mode of administrationand treatment. A wide array of formulations of the compounds andcompositions provided herein are contemplated as are a variety oftreatments for any disease or disorder described herein.

In human therapeutics, the physician will determine the dosage regimenthat is most appropriate according to a preventive or curative treatmentand according to the age, weight, stage of the disease and other factorsspecific to the subject to be treated. The compositions, in otherembodiments, should provide a dosage of from about 0.0001 mg to about 70mg of compound per kilogram of body weight per day. Dosage unit formsare prepared to provide from about 0.01 mg, 0.1 mg or 1 mg to about 500mg, or about 1000 mg, and in some embodiments from about 10 mg to about500 mg of the active ingredient or a combination of essentialingredients per dosage unit form.

The amount of active ingredient in the formulations provided herein,which will be effective in the prevention or treatment of a disorder orone or more symptoms thereof, will vary with the nature and severity ofthe disease or condition, and the route by which the active ingredientis administered. The frequency and dosage will also vary according tofactors specific for each subject depending on the specific therapy(e.g., therapeutic or prophylactic agents) administered, the severity ofthe disorder, disease, or condition, the route of administration, aswell as age, body, weight, response, and the past medical history of thesubject.

Exemplary doses of a formulation include milligram or microgram amountsof the active compound per kilogram of subject (e.g., from about 1microgram per kilogram to about 50 milligrams per kilogram, from about10 micrograms per kilogram to about 30 milligrams per kilogram, fromabout 100 micrograms per kilogram to about 10 milligrams per kilogram,or from about 100 microgram per kilogram to about 5 milligrams perkilogram).

It may be necessary to use dosages of the active ingredient outside theranges disclosed herein in some cases, as will be apparent to those ofordinary skill in the art. Furthermore, it is noted that the clinicianor treating physician will know how and when to interrupt, adjust, orterminate therapy in conjunction with subject response.

Different therapeutically effective amounts may be applicable fordifferent diseases and conditions, as will be readily known by those ofordinary skill in the art. Similarly, amounts sufficient to prevent,manage, treat or ameliorate such disorders, but insufficient to cause,or sufficient to reduce, adverse effects associated with the compositionprovided herein are also encompassed by the above-described dosageamounts and dose frequency schedules. Further, when a subject isadministered multiple dosages of a composition provided herein, not allof the dosages need be the same. For example, the dosage administered tothe subject may be increased to improve the prophylactic or therapeuticeffect of the composition or it may be decreased to reduce one or moreside effects that a particular subject is experiencing.

In certain embodiments, administration of the same formulation providedherein may be repeated and the administrations may be separated by atleast 1 day, 2 days, 3 days, 5 days, 10 days, 15 days, 30 days, 45 days,2 months, 75 days, 3 months, or 6 months.

BRIEF DESCRIPTION OF THE DRAWINGS

Embodiments of the invention are further described hereinafter withreference to the accompanying drawings, in which:

FIG. 1 shows an HPLC analysis of mesembrine;

FIG. 2 shows a ¹H NMR spectrum of mesembrine;

FIG. 3 shows a ¹H/¹³C HSQC NMR spectrum of mesembrine;

FIG. 4 shows an IR spectrum of mesembrine;

FIG. 5 shows an XRPD of damp (A), dry (B) and after 40° C./75% RH (C)solids from hydrochloric acid;

FIG. 6 shows a TG/DSC analysis of solids of hydrochloride salt Pattern1;

FIG. 7 shows a ¹H NMR spectrum of solids of hydrochloride salt pattern1, in d₆-DMSO;

FIG. 8 shows an example HPLC chromatogram of solids from the experimentwith hydrochloric acid, in ethanol;

FIG. 9 shows an XRPD of damp (A), dry (B) and after 40° C./75% RH solids(C) from benzenesulfonic acid;

FIG. 10 shows a TG/DSC analysis of solids of besylate salt Pattern 1;

FIG. 11 shows a ¹H NMR spectrum of solids of besylate salt pattern 1, ind₆-DMSO;

FIG. 12 shows an example HPLC chromatogram for solids of besylate saltpattern 1 from MEK;

FIG. 13 shows an XRPD of damp (A), dry (B) and after 40° C./75% RHsolids (C) from fumaric acid;

FIG. 14 shows a TG/DSC analysis of solids of fumarate salt Pattern 1(A), Pattern 2 (B) and Pattern 3 (C);

FIG. 15 shows a ¹H NMR spectrum of solids of fumarate salt Pattern 1(A), Pattern 2 (B) and Pattern 3 (C), in d₆-DMSO;

FIG. 16 shows an example HPLC chromatogram for solids of fumarate saltPattern 1 (A), Pattern 2 (B), Pattern 3 (C) from THF;

FIG. 17 shows a polarized light microscopy of hydrochloride salt Pattern1 (A), besylate salt Pattern 1 (B) and fumarate salt Pattern 1 and 4(C);

FIG. 18 shows an XRPD of solids from besylate salt Pattern 1 (A) andmagnified dried solids (B);

FIG. 19 shows an XRPD of lyophilized besylate salt;

FIG. 20 shows an XRPD of besylate salt Pattern 2;

FIG. 21 shows a comparison of the two crystalline besylate salt forms;

FIG. 22 shows the mean total concentrations of mesembrine besylate saltfollowing IP administration to male C57BI/6J mouse at 10.0 mg/kg; and

FIG. 23 shows the mean total concentrations of free base following IPadministration to male C57BI/6J mouse at 10.0 mg/kg.

DEFINITIONS

Various definitions are made throughout this document. Most words havethe meaning that would be attributed to those words by one skilled inthe art. Words specifically defined either below or elsewhere in thisdocument have the meaning provided in the context of the presentinvention as a whole and as typically understood by those skilled in theart.

The term “substantially crystalline” means at least about 50%crystalline and ranging up to about 100% crystalline. The presentinvention provides a salt that is at least about 50% crystalline, atleast about 60% crystalline, at least about 70% crystalline, at leastabout 80% crystalline, at least about 90% crystalline, at least about95% crystalline, at least about 98% crystalline, or at least about 100%crystalline in form.

The degree or percentage of crystallinity may be determined by theskilled person using X-ray powder diffraction (XRPD). Other techniques,such as solid-state nuclear magnetic resonance (NMR), FT-IR, Ramanspectroscopy, differential scanning calorimetry (DSC) andmicrocalorimetry, may also be used.

Crystalline forms of the salt of the invention may be in the form of asolvate, including but not limited to a hydrate (e.g., a monohydrate),or otherwise (e.g., in the form of an anhydrate).

“Subject,” “individual” or “patient” is used interchangeably herein andrefers to a vertebrate, preferably a mammal. Mammals include, but arenot limited to, murines, rodents, simians, humans, farm animals, sportanimals and pets.

“Treating” or “treatment” of any disease or disorder refers, in someembodiments, to ameliorating the disease or disorder (i.e., arresting orreducing the development of the disease or at least one of the clinicalsymptoms thereof,). Treatment may also be considered to includepreemptive or prophylactic administration to ameliorate, arrest orprevent the development of the disease or at least one of the clinicalsymptoms. Treatment can also refer to the lessening of the severityand/or the duration of one or more symptoms of a disease or disorder. Ina further feature, the treatment rendered has lower potential for longterm side effects over multiple years. In other embodiments “treating”or “treatment” refers to ameliorating at least one physical parameter,which may not be discernible by the patient. In yet other embodiments,“treating” or “treatment” refers to inhibiting the disease or disorder,either physically, (e.g., stabilization of a discernible symptom),physiologically, (e.g., stabilization of a physical parameter) or both.In yet other embodiments, “treating” or “treatment” refers to delayingthe onset of the disease or disorder.

“Therapeutically effective amount” means the amount of a compound that,when administered to a patient for treating a disease, is sufficient toeffect such treatment for the disease. The “therapeutically effectiveamount” will vary depending on the compound, the disease and itsseverity and the age, weight, adsorption, distribution, metabolism andexcretion etc., of the patient to be treated.

“Vehicle” refers to a diluent, excipient or carrier with which acompound is administered to a subject. In some embodiments, the vehicleis pharmaceutically acceptable.

“Active ingredient” or “Active pharmaceutical ingredient” or “API”refers to the novel mesembrine salt of the invention.

Mesembrine is a chiral alkaloid with the CAS name:(3aS-cis)-3a-(3,4-Dimethoxyphenyl)octahydro-1-methyl-6H-indol-6-one. Itoccurs naturally at the cis-isomer but may also be synthesized as thetrans-isomer or as a racemate. The structures below denote thestructural configuration of mesembrine. The mesembrine of the presentinvention may occur as the cis-isomer, the trans-isomer or a racemate ofthe two.

DETAILED DESCRIPTION OF THE INVENTION

The Examples below describe the phases of development of a highlysoluble and characterized mesembrine salt of the present invention. Theapplicant has determined that mesembrine is poorly soluble in aqueoussystems and as such means that the use of the compound as an activepharmaceutical ingredient in medicinal presentations is limited by itslack of solubility. A novel salt form has been produced which is bothsoluble and stable and as such is able to be used in the preparation ofmedicines and supplements.

Mesembrine can be isolated from extracts of S. tortuosum or can besynthesized chemically using the method described by Wang et al., 2016.

Example 1: Characterisation of Mesembrine Materials and Methods

The mesembrine used in the following series of experiments was isolatedfrom an extract of Sceletium tortuosum, the compound was characterizedby several different methods in order to determine the purity of thestarting material and to benchmark it against the subsequent salts thatwere to be formed.

HPLC analysis was carried out to determine the chemical purity of themesembrine and to provide information on any impurities. The followingconditions were use:

-   -   Column: Waters Acquity BEH C18 1.7 μm 150 mm×2.1 mm    -   Column Temperature: 33° C.    -   Autosampler Temperature: 25° C.    -   UV Wavelength: 280 nm    -   Injection Volume: 5 μL    -   Flow Rate: 0.29 mL/min    -   Mobile Phase A: 0.1% NH4 in H2O    -   Mobile Phase B: Methanol:Water 20:80% v/v    -   Gradient Program:

Time (minutes) Mobile Phase A [%] Mobile Phase B [%] 0.00 80 20 2.0 8020 4.0 60 40 6.0 50 50 9.0 50 50 14.0 10 90 17.0 10 90 17.1 80 20 23.080 20

¹H and ¹H/¹³C NMR analysis were carried out in order to determine thechemical structure of the mesembrine used. NMR experiments wereperformed on a Bruker AVIIIHD spectrometer equipped with a DCH cryoprobeoperating at 500.12 MHz for protons. Experiments were performed indeuterated DMSO, and each sample was prepared to ca. 10 mMconcentration.

The infrared (IR) spectrum was additionally recorded in order to obtaina benchmark of the starting mesembrine material. The followingconditions were used Infrared spectroscopy was carried out on a BrukerALPHA P spectrometer. Sufficient material was placed onto the centre ofthe plate of the spectrometer and the spectra were obtained using thefollowing parameters:

-   -   Resolution: 4 cm−1    -   Background Scan Time: 16 scans    -   Sample Scan Time: 16 scans    -   Data Collection: 4000 to 400 cm−1    -   Result Spectrum: Transmittance    -   Software: OPUS version 6

A Karl Fischer (KF) analysis was finally undertaken in order todetermine the water content of the mesembrine sample used. The followingconditions were used Solids were analysed either using a vaporizermethod, or using an external dissolution method:

Vaporizer method: Approximately 10 mg of material was weighted into a 10mL glass vial and tightly sealed with a screw cap. The water content ofthe samples was analysed using an InMotion KFoven Autosampler, at 150°C. The samples were run in duplicate, and an average moisture contentreported.

Blank Oven Temperature 150° C. Source for Drift Determination Max. StartDrift 10 μg/min Carrier Gas Flow Rate 80 mL/min Transfer Tube Heating NoMix Time 60 s Stir Speed 45% Drift Termination 10 s (Delay Time) Max.Titration Time 600 s Sample Oven Temperature 150° C. Source for DriftDetermination Max. start Drift 10 μg/min Carrier Gas Flow Rate 80 mL/minMix Time 60 s Stir Speed 45% Drift Termination 10 s (Delay Time) Max.Titration Time 600 s

External Dissolution method: A known mass of the sample to be analysedwas dissolved in a known mass of methanol. Prior to analysis beingcarried out, solvent blank measurements were carried out. Approximately1 mL of methanol was injected into the titration cell of a MettlerToledo C30 compact titrator, and the syringe back-weighed to determinethe weight of methanol added.

For the sample analysis, approximately 1 mL of the solution was injectedinto the titration cell, and the syringe back-weighed and the weight ofthe added solution entered onto the instrument. Titration was initiatedonce complete dissolution was observed, and the water content wasautomatically calculated by the instrument. The measurement was carriedout in duplicate and an average water content reported.

Results

FIG. 1 shows the HPLC analysis of the mesembrine starting material andthe impurity peaks found within the sample. It was determined that thepurity of the mesembrine was 91.17% are by HPLC, with 8.83% area by HPLCimpurities.

FIG. 2 details the ¹H NMR spectrum, it was determined that the spectrumwas consistent with the chemical structure of mesembrine. There was aresidual content of 5.0 wt % DCM which is equal to 0.2 equivalents ofDCM. No residual methanol was observed.

FIG. 3 details the ¹H/¹³C HSQC NMR spectrum. Analysis showed that thesample was consistent with mesembrine.

FIG. 4 details the IR spectrum of the mesembrine used and KF analysisdetermined that the water content of the received material was 1.1%(w/w).

Conclusion

The characterization of mesembrine determined that the sample used forsalt formation assessment was of an acceptable purity, matched theexpected chemical structure for mesembrine and comprised an acceptablelevel of water.

The mesembrine sample was therefore suitable for use in additionalexperiments to assess the ability of salt formation.

Example 2: Solubility of Mesembrine Materials and Methods

Approximately 360 mg of mesembrine was weighed out and dissolved in 5 mLof dichloromethane (DCM). After dissolution 278 μL of the solution wasaliquoted into pre-tared vials, and the solvent allowed to evaporate atambient conditions for 1 h. The vials were then dried at 20° C. undervacuum for ca. 22 h. After drying, the vials were back-weighed todetermine the mass of mesembrine in each vial.

The miscibility study was carried out by adding aliquots of the desiredsolvent system to each vial. The following aliquot sizes were used:Aliquot numbers 1-10—20 μL; 11-16—50 μL; 17-19—200 μL; 20—750 μL.

In between additions the experiment was heated to 40° C. for ca. 5minutes to facilitate miscibility/solubility. Additions were continueduntil miscibility was observed, or ca. 100 volumes (2 mL) had beenadded.

Results

Table 2.1 below details the approximate solvent miscibility/solubilityof mesembrine in the 18 different solvents tested.

TABLE 2.1 Solubility of mesembrine in various solvents # Solvent/% v/vMiscibility/mg/mL 1 2-Methyl THF >845 2 1,4-Dioxane >850 3 2-Propanol(IPA) >870 4 Acetone >890 5 Acetonitrile >855 6 2-Propanol:Water(50:50) >870 7 DMSO >288 8 Ethanol >630 9 Ethyl acetate >445 10 Heptane<9 11 Isopropyl acetate >900 12 Methanol >455 13 Methyl ethyl ketone(MEK) >915 14 N-Methyl-2-pyrrolidone (NMP) >450 15 Tert-butyl methylether >465 16 Tetrahydrofuran (THF) >381 17 Toluene >455 18 Water <9

As can be seen in Table 2.1 mesembrine was very poorly soluble inheptane and water and had a reasonable solubility in ethanol and otherorganic solvents.

Conclusion

Many organic solvents are classed as toxic or carcinogenic and as suchare not suitable for use as diluents in pharmaceutical compositions.

The low solubility of mesembrine in water means that the compound willbe difficult to formulate into a pharmaceutical composition as only asmall mass of the compound can be dissolved in water.

In a study determining the toxicity of Zembrin®, an extract of Sceletiumtortusosum, which comprises ca.20% mesembrine, the no observed adverseeffect level (NOAEL) was determined to be 420 mg extrapolated to a 70 kghuman (Murbach et al., 2014).

Therefore, a dose of around 84 mg of mesembrine (20% of 420 mg) wouldrequire approximately 10 ml of water to form a miscible solution, thisamount being far higher than the standard 00 capsule size used formedication delivery which has a fill volume of 0.9 ml.

Example 3: Primary Salt Formation Assessment Materials and Methods

The primary salt formation was assessed using reactions with 12counterions across six solvent systems. The various counterions wereselected based on their pKa, and also as a result of their molecularweight, toxicity and diversity. The counterions and solvents used forthe primary salt formation assessment are as detailed in Table 3.1below.

TABLE 3.1 Counterions and solvent details Upper Merck Solvent Temp. #Counterions MW/g/mol pKa Class System/% v/v Limit/º C. 1 Hydrochloricacid 36.46 −6 1 THF 40 2 Sulfuric acid 98.08 −3 1 Ethyl acetate 3p-Toluenesulfonic acid 172.2 −1.34 2 Ethanol 4 Methanesulfonic acid96.10 −1.2 2 IPA:Water 50:50 5 Benzenesulfonic acid 158.18 0.7 2 MEK 6Maleic acid 116.08 1.92; 6.23 1 Acetonitrile 7 Phosphoric acid 98.001.96; 7.12; 12.32 1 8 (+)-L-Tartaric acid 150.09 3.02; 4.36 1 9 Fumaricacid 116.08 3.03; 4.38 1 25 10 Citric acid 192.13 3.13; 4.76; 6.40 1 11Succinic acid 118.09 4.21; 5.64 1 12 Benzoic acid 179.18 4.3 3

Stock solutions of mesembrine at 200 mg/mL were prepared in the desiredsolvent systems. Separately, 1.1 equivalents of counterion weredispensed (either weighed, or measured using an autopipette) into 72×2mL vials and a stirrer bar added.

100 μL of the appropriate solvent system was added to each vial todissolve/suspend the counterion. 100 μL of the mesembrine stock solutionin the correct solvent system was added to each vial, at the specifiedupper temperature limit.

The experiments were then temperature cycled between the uppertemperature limit (specified in Table 3.1) and 5° C. at 0.1° C./min,with 1 h holds at the upper temperature limit, and 5° C. for ca. 72 h.

After temperature cycling, solids were isolated from experiments at 5°C., and analysed by XRPD. Any experiments which did not contain solidshad anti-solvent addition carried out and were temperature cycled for afurther ca. 24 h as above.

Solids were dried at 40° C. under vacuum for ca. 24 h, and thenre-analysed by XRPD.

Solids were then stored at 40° C./75% RH for ca. 24 h, and thenre-analysed by XRPD.

Potential salt forms were also analysed by TGA/DSC.

Additional experiments were carried out for selected counterions(numbers 2, 3, 4 and 6 from Table 3.1) had not yielded any solids of apotential salt of mesembrine from the initial set of salt formations.The upper temperature limit was decreased from 40° C. to 25° C. asdegradation was noted in these during the initial experiments asdetailed below and in Table 3.2.

A stock solution of mesembrine at 200 mg/mL was prepared in MEK.Separately, 1.1 equivalents of counterion were dispensed (eitherweighed, or measured using an autopipette) into 4×2 mL vials and astirrer bar added.

100 μL of MEK was added to each vial to dissolve/suspend the counterion.100 μL of the mesembrine stock solution was added to each vial, at 25°C.

The experiments were then temperature cycled between 25° C. and 5° C. at0.1° C./min, with 1 h holds at 25° C. and 5° C. for ca. 72 h. Aftertemperature cycling, anti-solvent addition carried out in 100 μLaliquots, at 5° C. until a visual change was observed or 1 mL of heptanehad been added.

The experiments were temperature cycled for a further ca. 24 h as above.

TABLE 3.2 Counterions and solvent details for additional salt formationassessment Solvent Upper MW/ Merck System/ Temp. # Counterions g/mol pKaClass % v/v Limit/º C. 1 p-Toluenesulfonic 172.2 −1.34 2 MEK 25 acid 2Methanesulfonic 96.10 −1.2 2 acid 3 Maleic acid 116.08 1.92; 6.23 1 4Sulfuric acid 98.08 −3 1

Results

The primary salt formation assessment from reactions with 12 counterionsacross six different solvent systems were determined. The results of theprimary salt screen are summarized in Tables 3.3 to 3.5 below.

TABLE 3.3 Summary of damp XRPD analysis from primary formationassessment Ethyl IPA:Water Counterions THF Acetate Ethanol 50:50% v/vMEK MeCN Hydrochloric acid 1, C 1, PC 1, C — 1, C 1, C Sulfuric acid — —— — — — p-Toluenesulfonic acid — — — — — — Methanesulfonic acid — — — —— — Benzenesulfonic acid 1, C 1, C 1, C — 1, C 1, C Maleic acid — — — —— — Phosphoric acid 1, PC A 2, PC — 1, PC — (+)-L-Tartaric acid A U 1,PC — 1, PC 1, C Fumaric acid 1, PC 2, C 2, C — 1, PC 3, C Citric acid AU — — A A Succinic acid — U — — 1, C — Benzoic acid — — — — — — Key AAmorphous # Pattern number C Crystalline potential salt PC Poorlycrystalline U Counterion (unreacted) — No solids

TABLE 3.4 Summary of dry XRPD analysis from primary formation assessmentEthyl IPA:Water Counterions THF Acetate Ethanol 50:50% v/v MEK MeCNHydrochloric acid 1, C 1, PC 1, C — 1, C 1, C Sulfuric acid — — — — — —p-Toluenesulfonic acid — — — — — — Methanesulfonic acid — — — — — —Benzenesulfonic acid 1, C 1, C 1, C — 1, C 1, C Maleic acid — — — — — —Phosphoric acid 1, PC A 2, PC — 1, PC — (+)-L-Tartaric acid 1, PC U 1,PC — 1, PC 1, C Fumaric acid 1, PC 2, C 2, C — 1, PC 3, C Citric acid AU — — A A Succinic acid — U — — 1, C — Benzoic acid — — — — — — Key AAmorphous # Pattern number C Crystalline potential salt PC Poorlycrystalline U Counterion (unreacted) — No solids

TABLE 3.5 Summary of 40° C./75% RH XRPD analysis from primary formationassessment Ethyl IPA:Water Counterions THF Acetate Ethanol 50:50% v/vMEK MeCN Hydrochloric acid 1, C A 1, C — 1, C 1, C Sulfuric acid — — — —— — p-Toluenesulfonic acid — — — — — — Methanesulfonic acid — — — — — —Benzenesulfonic acid 1, C 1, C 1, C — 1, C 1, PC Maleic acid — — — — — —Phosphoric acid 3, C 3, C 3, C — 3, C — (+)-L-Tartaric acid A A A — A 1,PC Fumaric acid 1, C 1, C 1*, C — ¼, C ¼, C Citric acid A A — — A ASuccinic acid — U — — A — Benzoic acid — — — — — — Key A Amorphous #Pattern number C Crystalline potential salt PC Poorly crystalline UCounterion (unreacted) — No solids * Additional peaks

A summary of the properties of the hydrochloride, besylate and fumaratesalts are reported in Table 3.6 below in addition to FIGS. 5 to 16 .

TABLE 3.6 Summary of salt properties TGA/DSC Weight Loss/ EndothermicExothermic HPLC wt % Events Events Solid (Temp/ (onset/peak)/(onset/peak)/ Purity/ º C.) º C. º C. % area ¹H NMR CommentsHydrochloride Pattern 2.1 194/208 N/A 92.2 Ethanol 0.5 Anhydrous, high 1(20-191) wt % potential melt, physically stable at 40° C./75% RH,solution degradation observed Besylate Pattern 1 1.1 138/149 N/A 91.5Benzenesulfonic Anhydrous, (20-196) acid 1.3 equiv. physically stableEthanol 0.37 at 40° C./75% RH, wt % solution degradation observedPhosphate Pattern 1 10.5 82/113 N/A 93.4 THF 2.3 wt % Likely solvate.(20-211) Physically unstable at 40° C./75% RH. Solution degradationobserved. Phosphate Pattern 2 9.0 N/A N/A 92.7 Ethanol 1.6 Potential(20-201) wt % solvate/hydrate. Physically unstable at 40° C./75% RHPhosphate Pattern 3 1.7 103/113 167/171 N/A No residual Obtained from(20-119) 218/230 solvents exposure to 5.3 40° C./75% RH, (119-199)multiple weight 7.5 losses in TGA. (199- 300 Tartrate Pattern 1 2.3126/141 N/A 91.7 Tartrate 1.1 Potentially hydrate (20-164) 171/192equiv. or anhydrous. 35.9 No residual Deliquescence (164- solventsobserved at 301) 40° C./75% RH. Solution degradation observed. FumaratePattern 1 4.3 143/153 N/A 90.4 Fumarate 1.5 Likely solvate. (20-168)equivalents Physically stable 31.2 THF 1.7 wt % at 40° C./75% RH, (168-from THF. 336) Fumarate Pattern 2 5.7 141/152 N/A 94.4 Fumarate 1.3Likely solvate. (72-159) 200/205 equivalents Physically 31.1 Ethanol 2.9unstable at (159- wt % 40° C./75% RH. 330) Solution degradationobserved. Fumarate Pattern 3 2.9 139/147 N/A 96.9 Fumarate 1.4Potentially (112-175) 186/207 equivalents anhydrous or 39.1 No residualhydrate. Physically (175- solvents unstable at 304) 40° C./75% RH.Fumarate Pattern 1/4 7.7 137/148 N/A 93.1 Fumarate 1.2 Obtained from(20-170) equivalents exposure to 30.6 No residual 40° C./75% RH. (170-solvents 303) Succinate Pattern 1 2.5 74/83 N/A 89.2 Succinate 1.9Potentially (20-127) 152/161 equivalents anhydrous or 56.0 No residualhydrate. Low (127- solvents potential melt. 316) Physically unstable at40° C./75% RH. Solution degradation observed.

As can be seen from the tables above no solids were isolated for thefollowing counterions: sulfuric acid, p-toluenesulfonic acid,methanesulfonic acid, malic acid and benzoic acid. Additionalexperiments were performed on these counterions as described in themethodology section, however after temperature cycling and anti-solventaddition, no solids were observed.

The results from reactions with each acid where solids were formed aresummarised in more detail below.

Hydrochloric Acid:

When hydrochloric acid was used, the following results and observationswere obtained.

Solids of potential hydrochloride salt Pattern 1 were isolated from fivesolvent systems: THF, ethyl acetate, ethanol, MEK and acetonitrile. XRPDanalysis indicated that hydrochloride salt Pattern 1 persisted afterdrying at 40° C. under vacuum and after storage at 40° C./75% RH asshown in FIG. 5 .

TG/DSC analysis of the solids of hydrochloride salt Pattern 1, isolatedfrom ethyl acetate, showed a weight loss of 2.1 wt % between 20-191° C.One endothermic event was observed with an onset temperature of 194° C.,and a peak temperature of 208° C. as shown in FIG. 6 .

The ¹H NMR spectrum of the solids of hydrochloride salt Pattern 1,isolated from ethanol, was consistent with the chemical structure ofmesembrine, with an additional signal at 11.39 ppm, consistent with saltformation. The residual ethanol content was 0.5% wt as shown in FIG. 7 .

HPLC analysis on the dried solids from ethanol showed the chemicalpurity was 92.2% area (c.f.: the input purity was 91.2% area) as shownin FIG. 8 .

HPLC analysis on the filtered mother liquor from the experiment in ethylacetate indicated the chemical purity was 40.3% area.

Benzenesulfonic Acid:

The results of the experiments with benzenesulfonic acid are summarizedbelow.

Solids of potential besylate salt Pattern 1 were isolated from fivesolvent systems: THF, ethyl acetate, ethanol, MEK and acetonitrile. ThisXRPD pattern persisted after drying at 40° C. under vacuum, and afterstorage at 40° C./75% RH as shown in FIG. 9 .

TG/DSC analysis of dried solids of besylate salt Pattern 1, isolatedfrom ethyl acetate, showed a weight loss of 1.1 wt % between 20-196° C.In the DSC thermogram, one endothermic event was observed with an onsettemperature of 138° C., and a peak temperature of 149° C. as shown inFIG. 10 .

The ¹H NMR spectrum of solids of besylate salt Pattern 1, isolated fromethanol, was consistent with the structure of mesembrine, with anadditional signal at 9.92 ppm, indicative with salt formation.Benzenesulfonic acid was also observed in the NMR spectrum, and thecontent was equal to 1.3 equivalents. A residual ethanol content of 0.37wt % was observed as shown in FIG. 11 .

HPLC analysis of the dried solids from the experiment in MEK indicated achemical purity of 91.46% area (c.f.: the input purity was 91.2% area)as shown in FIG. 12 .

HPLC analysis of the mother liquor from the experiment in ethyl acetateindicated a chemical purity of 44.21% area.

Phosphoric Acid:

The results of the experiments with phosphoric acid are summarizedbelow.

Solids isolated from THF and MEK were poorly crystalline and labelledpotential phosphate salt Pattern 1. After drying at 40° C. under vacuumsolids of poorly crystalline phosphate Pattern 1 were maintained. Afterstorage at 40° C./75% RH, the solids isolated from MEK had deliquescedbut re-solidified out of the high humidity. For both materials,conversion to phosphate salt Pattern 3 was observed.

The solids isolated from the experiment in ethyl acetate were amorphousand remained amorphous after drying. After 24 h at 40° C./75% RH,conversion to phosphate Pattern 3 was observed.

The solids isolated from ethanol were poorly crystalline and labelled aspotential phosphate salt Pattern 2. After drying at 40° C. under vacuumsolids of poorly crystalline phosphate Pattern 2 were maintained, andafter storage at 40° C./75% RH, conversion to phosphate salt Pattern 3was observed.

HPLC analysis of the dried solids of phosphate salt Pattern 1, from THF,showed a chemical purity of 93.4% area. HPLC analysis of dried solids ofphosphate salt Pattern 2, from ethanol, showed a chemical purity of92.67% area.

The purity of the mother liquor from the experiment in MEK was 65.31%area.

TG/DSC analysis of solids of phosphate salt Pattern 1, from THF, showeda weight loss of 10.5 wt % between 20-211° C. In the DSC thermogram, oneendothermic event was observed with an onset temperature of 82° C., anda peak temperature of 113° C.

The ¹H NMR spectrum of phosphate salt Pattern 1, isolated from THF, wasconsistent with the structure of mesembrine. A residual THF content of2.3 wt % was observed. The 31P NMR analysis indicated a phosphate groupwas present in the material.

The TG/DSC analysis of solids of phosphate salt Pattern 2, from ethanol,showed a weight loss of 9.0 wt % between 20-201° C. In the DSCthermogram, no endothermic events were observed.

The ¹H NMR spectrum of phosphate salt Pattern 2, isolated from ethanol,was consistent with the structure of mesembrine. A residual ethanolcontent of 1.6 wt % was observed. The 31P NMR spectrum was consistentwith the presence of a phosphate group within this material.

The TG/DSC analysis of solids of phosphate salt Pattern 3, obtained fromTHF after storage at 40° C./75% RH, showed a weight loss of 1.7 wt %between 20-119° C., followed by a second weight loss of 5.3 wt % between119-199° C. a final weight loss of 7.5 wt % was observed between199-300. In the DSC thermogram there was one endothermic event with anonset temperature of 103° C. and a peak temperature of 113° C. In theDSC thermogram there were also two exothermic events with onsettemperatures of 167° C. and 218° C., and peak temperatures of 171° C.and 230° C.

The ¹H NMR spectrum of phosphate salt Pattern 3, isolated from MEK afterstorage at 40° C./75% RH was consistent with the structure ofmesembrine. No residual solvent was detected. The 31P NMR spectrumindicated the presence of a phosphate group within the material.

(+)-L-Tartaric Acid:

The results of the experiments with (+)-L-tartaric acid are summarizedbelow.

Solids of potential tartrate salt, Pattern 1, were isolated from ethanoland MEK, but were poorly crystalline, and from acetonitrile. Afterdrying under vacuum, solids of tartrate salt Pattern 1 were maintainedin solids from ethanol, MEK and acetonitrile. After exposing to 40 C/75%RH, the solids from ethanol and MEK had deliquesced and re-solidified onremoval from the humidity chamber. These solids were amorphous. Thesolids from acetonitrile remained as tartrate salt Pattern 1, but with adecrease in crystallinity.

Solids of tartrate salt Pattern 1 were also observed in the solidsisolated from THF, after drying at 40° C. under vacuum.

The solids of tartrate salt Pattern 1, isolated from THF, had a chemicalpurity of 91.70% area (c.f.: the input purity was 91.2% area) by HPLC.

The mother liquor for the experiment in acetonitrile had a chemicalpurity of 34.80% area.

TG/DSC analysis of solids of tartrate salt Pattern 1, from ethanol,showed a weight loss of 2.3 wt % between 20-164° C., followed by asecond weight loss of 35.9 wt % between 164-301° C. Two endothermicevents were observed in the DSC thermogram with onset temperatures of126° C. and 171° C., and peak temperatures of 141° C. and 192° C.

The 1H NMR spectrum of solids of tartrate salt Pattern 1, isolated fromacetonitrile, was consistent with the structure of mesembrine. Tartaricacid was observed, with a content equal to 1.1 equivalents. No residualacetonitrile was observed.

Fumaric Acid:

The results of the experiments with fumaric acid are summarized below.

Solids isolated from THF were poorly crystalline and labelled potentialfumarate salt Pattern 1. After drying at 40° C. under vacuum this poorlycrystalline Pattern 1 material persisted. After storage at 40° C./75%RH, an increase in crystallinity was observed, and the material remainedPattern 1.

Solids isolated from ethyl acetate were crystalline and labelled aspotential fumarate salt Pattern 2. After drying at 40° C. under vacuum amixture of fumarate Pattern 1 and Pattern 2 was observed by XRPD. Afterstorage at 40° C./75% RH, conversion to fumarate salt Pattern 1 wasobserved.

Solids of fumarate salt Pattern 2 were also observed from ethanol, whichpersisted after drying. After exposure to 40° C./75% RH conversion tofumarate salt Pattern 1, with additional peaks, was observed.

Solids of poorly crystalline fumarate salt Pattern 1 were observed fromMEK, and were maintained after drying at 40° C. under vacuum. Exposureto 40° C./75% RH resulted in conversion to a mixture of fumarate saltPattern 1 and Pattern 4.

Solids isolated from acetonitrile were crystalline, and labelled asfumarate salt Pattern 3, and persisted after drying at 40° C. undervacuum. Exposure to 40° C./75% RH resulted in conversion to a mixture offumarate salt Pattern 1 and Pattern 4.

XRPD of the salt Patterns 1, 2 and 3 are shown in FIG. 13 .

TG/DSC analysis showed the solids of fumarate salt Pattern 1, isolatedfrom THF, showed a weight loss of 4.3 wt % between 20-168° C., followedby a second weight loss of 31.2 wt % between 168-336° C. One endothermicevent was observed in the DSC thermogram with an onset temperature of143° C. and a peak temperature of 153° C.

The ¹H NMR spectrum of fumarate salt Pattern 1, isolated from THF, wasconsistent with the structure of mesembrine. The fumaric acid contentwas equal to 1.5 equivalents (30.7 wt %). The residual THF content was1.7 wt %.

TG/DSC analysis showed the solids of fumarate salt Pattern 2, isolatedfrom ethanol, showed a weight loss of 5.7 wt % between 72-159° C., and asecond weight loss of 31.1 wt % between 159-330° C. There were twoendothermic events in the DSC thermogram with onset temperatures of 141°C., and 200° C., and peak temperatures of 152° C. and 205° C.

The ¹H NMR spectrum of fumarate salt Pattern 2, isolated from ethanol,was consistent with the structure of mesembrine. The fumaric acidcontent was equal to 1.3 equivalents (27.6 wt %). The residual ethanolcontent was 2.9 wt %.

TG/DSC analysis of solids of fumarate salt Pattern 3, isolated fromacetonitrile, showed a weight loss of 2.9 wt % between 112-175° C.,followed by a second weight loss of 39.1 wt % between 175-304° C. Twoendothermic events were observed with onset temperatures of 139° C. and186° C., and peak temperatures of 147° C. and 207° C.

The ¹H NMR spectrum of fumarate salt Pattern 3, isolated fromacetonitrile, was consistent with the structure of mesembrine. Thefumaric acid content was equal to 1.4 equivalents. No residualacetonitrile was observed.

The TG/DSC analysis of solids of a mixture of fumarate salt Pattern 1and Pattern 4, isolated from acetonitrile after storage at 40° C./75%RH, showed a weight loss of 7.7 wt % between 20-170° C., and a secondweight loss of 30.6 wt % between 170-303° C. In the DSC thermogram, oneendothermic event was observed with an onset temperature of 137° C. anda peak temperature of 148° C.

The ¹H NMR spectrum of a mixture of fumarate salt Pattern 1+Pattern 4,isolated from MEK after exposure to 40° C./75% RH, was consistent withthe chemical structure of mesembrine. The fumaric acid content was equalto 1.2 equivalents. No residual MEK was observed.

The TG/DSC for salt Patterns 1-3 are shown in FIG. 14 and the ¹H NMRspectra for salt Patterns 1-3 are shown in FIG. 15 .

HPLC analysis showed a chemical purity of 90.36% area for the solids offumarate salt Pattern 1, isolated from THF. The solids of fumarate saltPattern 2, isolated from ethanol, had a chemical purity of 94.35% area.The solids of fumarate salt Pattern 3, isolated from acetonitrile, had achemical purity of 96.47% area. The solids of a mixture of fumarate saltPattern 1 and Pattern 4, isolated from MEK after exposure to 40° C./75%RH, was 93.1% area c.f.: the input purity was 91.2% area) as shown inFIG. 16 .

Succinic Acid:

The results of the experiments with succinic acid are summarized below.

The solids isolated from MEK were crystalline and labelled potentialsuccinate salt Pattern 1. This pattern persisted after drying at 40° C.under vacuum, however after exposure to 40° C./75% RH, conversion toamorphous material was observed.

Solids of potential succinate salt Pattern 1 had a chemical purity of89.2% area.

The TG/DSC analysis of potential succinate salt Pattern 1 showed aweight loss of 2.5 wt % between 20-127° C., followed by a second weightloss of 56.0 wt % between 127-316° C. In the DSC thermogram, twoendothermic events were observed with onset (and peak) temperatures of74° C. (83° C.), and 152° C. (161° C.).

The ¹H NMR analysis of potential succinate salt Pattern 1 was consistentwith the chemical structure of mesembrine. A succinic acid content of35.9 wt % (1.9 equivalents) was observed. No residual solvents weredetected.

Conclusion

The data detailed above demonstrates that the salts formed using thecounterions hydrochloric acid, benzenesulfonic acid and fumaric acidwere crystalline, anhydrous solids with high potential melt temperaturesand able to demonstrate physical stability.

As such these counterions were selected for a secondary salt formationassessment.

Table 3.1 details the pKa of the various counterions tested in the saltformation assessment. It is generally accepted that a difference of atleast 2 pKa units between the acid and base are required for protontransfer, and as such the stronger the acid (lower pKa) the more likelya salt is to form. It was therefore surprising that apart fromhydrochloric acid the stronger acids such as sulfuric acid,p-toluenesulfonic acid, methanesulfonic acid were unable to form saltswith mesembrine.

Example 4: Secondary Salt Formation Assessment Materials and MethodsHydrochloride Salt Pattern 1:

The following procedure was used for the scale-up of the Hydrochloridesalt Pattern 1. Approximately 0.5 g of mesembrine was dissolved in 2.5mL of ethanol. 1.1 equivalents of HCl (0.156 mL) were dissolved in 2.5mL of ethanol, and added to the solution of mesembrine, at 25° C.(Initial concentration: 100 mg/mL).

The experiment was stirred for ca. 1 h, then cooled to 5° C. at 0.1°C./min. After ca. 18 h at 5° C., 200 μL was sub-sampled and centrifugedand the solids analysed by XRPD.

The solids were predominately amorphous, so the experiment was heated to25° C. over 10 minutes, and 15 mL of heptane was added over 30 minutes.The experiment was then cooled to 5° C. at 0.1° C./minute and held at 5°C. After ca. 2 h at 5° C., 200 μL was sub-sampled and centrifuged andthe solids analysed by XRPD. The mother liquor was biphasic.

The solids were poorly crystalline, so the experiment was temperaturecycled between 25° C. and 5° C. at 0.1° C./min, with 1 h holds at 25° C.and 5° C., for ca. 18 h. At 5° C., the solids were isolated by Buchnerfiltration and analysed by XRPD. The solids were dried at 40° C. undervacuum for ca. 72 h and then further characterization carried out.

Besylate Salt Pattern 1:

The following procedure was used for the scale-up of the Besylate saltPattern 1.

Approximately 0.5 g of mesembrine was dissolved in 2.5 mL of MEK. 1.1equivalents of benzenesulfonic acid (306.78 mg) were dissolved in 2.5 mLof MEK, and added to the solution of mesembrine, at 25° C.(Concentration: 100 mg/mL).

The experiment was stirred for ca. 1.5 h. The experiment was then cooledto 5° C. at 0.1° C./minute and held at 5° C. for ca. 18 h. A sub-sample(200 μL) was taken and centrifuged, and the solids analysed by XRPD.

The solids were isolated by Buchner filtration and dried at 40° C. undervacuum for ca. 18 h before being characterized further.

Fumarate Salt Pattern 1+Pattern 4 (Mixture):

The following procedure was used for the scale-up of the fumarate saltPattern 4.

Approximately 0.5 g of mesembrine was dissolved in 2.5 mL of MEK. 1.1equivalents of fumaric acid (222 mg) were dissolved in 2.5 mL of MEK,and the solution of mesembrine was added to the fumaric acid, dropwiseat 25° C. (Concentration: 100 mg/mL).

The experiment was stirred for ca. 1 h at 25° C. The experiment was thencooled to 5° C. at 0.1° C./min and held at 5° C. for ca. 18 h. Asub-sample (200 μL) was taken and centrifuged, and the solids analysedby XRPD.

The slurry was very thick, therefore an additional 2 mL of MEK was addedto improve slurry mobility, and stirred for ca. 30 min. The solids wereisolated by Buchner filtration and dried at 40° C. under vacuum for ca.18 h. The solids were analysed by XRPD, and then transferred to 40°C./75% RH.

After 5 days at 40° C./75% RH the solids were removed and furthercharacterized.

The solids were sub-sampled after 1, 2 and 5 days for XRPD analysis.

Re-Preparation of Fumarate Salt Pattern 4:

The following general procedure was used for the re-preparation of thefumarate salt Pattern 4.

Approximately 103 mg of mesembrine was weighed into a vial and dissolved0.52 mL of MEK at 25° C. 1.1 equivalents of fumaric acid was suspendedin 0.52 mL of MEK. The mesembrine solutions were added to the fumaricacid at 25° C. and stirred.

The experiments were stirred for 1.5 h at 25° C. and then was cooled to5° C. and 0.1° C./min and held at 5° C. for ca. 18 h before isolation.The solids were isolated by centrifugation. Solids were isolated anddried at 40° C. under vacuum for ca. 24 h.

Results Hydrochloride Salt Pattern 1:

Solids of hydrochloride salt Pattern 1 were isolated in a 48% yield,with a chemical purity of 90.5% area. KF indicated a water content of0.4% w/w. CAD indicated a chloride content of 0.9 equivalents,indicating the material is a mono-hydrochloride salt.

The solids were faint beige (BE10) with respect to the Sigma Aldrichcolour chart.

TG/DSC analysis showed a weight loss of 1.7 wt % between 20-160° C.,followed by a weight loss of 21.7 wt % between 162-273° C. Anendothermic event with an onset temperature of 188° C. and a peaktemperature of 200° C. was observed.

DSC analysis showed an endothermic event with an onset temperature of182° C. and a peak temperature of 197° C.

VT-XRPD analysis showed that solids of hydrochloride salt Pattern 1persisted between 25-160° C. At 188 and above (to 205° C.), the materialwas amorphous, and remained amorphous after cooling. The solids appearedto have melted.

Polarized light microscopy (PLM) showed the material had no distinctmorphology, with no apparent birefringence, see FIG. 17A.

An IR spectrum was recorded for reference.

The ¹H NMR spectrum was consistent with the formation of a salt ofmesembrine. A residual ethanol content of 0.98 wt %, and a residualheptane content of 0.64 wt % was observed.

DVS analysis showed a water uptake of 5.4 wt % at 80% RH, and 25° C.,indicating the material was hygroscopic. At 90% RH, the water uptake was20.3 wt % indicating significant hygroscopicity above 80% RH. Solids ofhydrochloride salt Pattern 1 were recovered from the DVS analysis.

VH-XRPD analysis showed that even after 12 h at 90% RH, the solidsremained consistent with hydrochloride salt Pattern 1. No change incrystallinity was observed.

Besylate Salt Pattern 1:

Solids of besylate salt Pattern 1 were isolated in a 69% yield, with achemical purity of 98.1% area. KF indicated a water content of 0.3% w/w.

The solids were faint beige (BE10) with respect to the Sigma Aldrichcolour chart.

TG/DSC analysis showed a weight loss of 0.3 wt % prior to decomposition.In the DSC thermogram two endothermic events were observed with onset(and peak) temperatures of 131° C. (141° C.) and 151° C. (158° C.).

In the DSC, at a heating rate of 10° C./min, two overlapping endothermicevents were observed in the first heat with onset (and peak)temperatures of 132° C. (141° C.) and 150° C. (158° C.). In the firstcool a vitrification was observed with a mid-point half-height of 59°C., in the second heat a glass transition was observed with a mid-pointhalf-height of 65° C. The DSC was repeated with a heating rate of 1°C./min which successfully resolved the endothermic events in the firstheat, which were observed to have onset (and peak) temperatures of 129°C. (135° C.), and 151° C. (156° C.).

VT-XRPD analysis showed that solids of besylate salt Pattern 1 persistedbetween 25-150° C. At temperatures of 157° C. to 160° C., and afterreturning to 25° C., the material was amorphous, and appeared to havemelted.

PLM indicated the material had no distinct morphology, with somebirefringence, see FIG. 17B.

An IR spectrum was recorded for reference.

The ¹H NMR spectrum was consistent with the formation of a salt ofmesembrine. The benzenesulfonic acid content was 1.1 equivalents,indicating the material is a mono-besylate salt.

DVS analysis showed that besylate salt Pattern 1 had a water uptake of2.3 wt % at 80% RH and 25° C., indicating the material was hygroscopic.At 90% RH, the water uptake was 13 wt %, indicating significanthygroscopicity above 80% RH. Solids isolated at the end of the DVSanalysis were consistent with besylate salt Pattern 1.

VH-XRPD analysis showed that even after 17 h at 90% RH, solids ofbesylate salt Pattern 1 persisted. There was no change in crystallinityduring this analysis.

Fumarate Salt Pattern 1+Pattern 4 (Mixture):

Solids of a mixture of fumarate salt Pattern 1 and Pattern 4 wereisolated in a 55% yield, with a chemical purity of 96.6% area. KFanalysis indicated a water content of 4.0% w/w (ca. 1 equivalent water).

The solids were faint beige (BE10) with respect to the Sigma Aldrichcolour chart.

TG/DSC analysis showed a weight loss of 3.4 wt % between 20-95° C.,followed by second weight loss of 1.8 wt % between 95° C. and 162° C. Inthe DSC thermogram, two endothermic events were observed with onset (andpeak) temperatures of 67° C. (80° C.), and 131° C. (154° C.).

In the DSC thermogram, two endothermic events were observed with onset(and peak) temperatures of 61° C. (75° C.), and 136° C. (152° C.).

VT-XRPD showed that between 25° C. and 90° C. a mixture of fumarate saltPattern 1 and Pattern 4 persisted. At 130° C., the diffractogram wasconsistent with fumarate Pattern 4. At 154° C. and to 170° C., and afterreturning to ambient temperature the material was amorphous.

PLM showed the material had no distinct morphology, with limitedbirefringence, see FIG. 17C.

An IR spectrum was recorded for reference.

The ¹H NMR spectrum was consistent with the salt formation ofmesembrine. The fumaric acid content was equal to 1.3 equivalents,indicating this material is a mono-fumarate salt. A residual MEK contentof 0.74 wt % was observed.

DVS analysis showed that at 80% RH and 25° C., the water uptake was 0.7wt %, indicating that the material is slightly hygroscopic. Between0-20% RH, a water uptake of 2.1 wt % was observed, indicating that atambient humidity's the material is likely hydrated. The solids isolatedat the end of the analysis were consistent with a mixture of fumaratesalt Pattern 1 and Pattern 4.

VH-XRPD showed that between 40% RH and 10% RH, the solids wereconsistent with a mixture of fumarate salt Pattern 1 and Pattern 4. At0% RH, a new diffractogram was observed, labelled fumarate Pattern 5.

Re-Preparation of Fumarate Salt Pattern 4:

XRPD showed Pattern 4 without Pattern 1 present. DVS analysis onfumarate salt Pattern 4 showed a water uptake of 0.5 wt % at 80% RH, 25°C., indicating the material was slightly hygroscopic. HPLC indicated thechemical purity was 95.88% area.

Conclusion

Salts were successfully formed from all three counterions and analysedaccordingly. The three salts demonstrated properties consistent withimproved attributes consistent with the ability to produce enhancedpharmaceutical formulations.

Example 5: Salt Hydration Study Materials and Methods

Approximately 20 mg of the hydrochloride (HCl), besylate and fumaratesalts were weighed into 3×2 mL push cap vials. 0.05-0.1 mL of relevantsolvent system was added to each vial, at 25° C.

Methanol/water solvent systems (99:1, 75:25, 33:67) of known wateractivity (a_(w) 0.2, 0.5 and 0.8 respectively) were used. The experimentwas stirred for ca. 24 h. After 24 h, any slurries were centrifuged, andthe solids analysed by XRPD.

An additional hydration study was required in order to repeat the salthydration studies, for the HCl salt and the besylate salt due todissolution in the first set of experiments:

For each salt, approximately 10 mg was weighed into 4×2 mL vials and astirrer bar added. 10 μL of each solvent system was added, at 25° C. Theexperiments were stirred for 24 h, and any solids recovered were thenanalysed by XRPD.

Results

The results of the hydration studies are summarized in Table 3.7 below.

Clear solutions were obtained for the experiments with hydrochloridesalt Pattern 1, and besylate salt Pattern 1.

Additional experiments with hydrochloride salt Pattern 1 resulted inslurries persisting at a_(w) 0.5 and 0.8. The isolated solids wereconsistent with hydrochloride salt Pattern 1.

Clear solutions were obtained from the additional experiments withbesylate salt Pattern 1, indicating the solubility was >1000 mg/mL.

Slurries were maintained for the fumarate salt. The isolated solids wereconsistent with fumarate salt Pattern 4.

TABLE 3.7 Summary of Results from Hydration Studies SaltConcentration/mg/mL Water Activity/a_(w) Observation XRPD Hydrochloride200 0.2 Clear solution — Pattern 1 1000 — 200 0.5 Clear solution — 1000Slurry 1, C 200 0.8 Clear solution 1000 Slurry 1, C Besylate 200 0.2Clear solution — Pattern 1 1000 400 0.5 Clear solution — 1000 400 0.8Clear solution — 1000 Fumarate 400 0.2 Thick slurry 4, C Pattern 1 + 4400 0.5 Slurry 4, C 200 0.8 Thick slurry 4, C Key A Amorphous # Patternnumber C Crystalline potential salt PC Poorly crystalline U Counterion(unreacted) — No solids

Conclusion

The besylate salt was highly soluble resulting in greater than 1000mg/ml at all levels of water activity, providing evidence that this saltis highly suitable for development of a pharmaceutical composition.

The hydrochloride salt was not soluble at higher concentrations andhigher water activity levels. The fumarate did not produce a clearsolution at any of the concentrations or water activity levels tested.

Example 6: PH Solubility Study

Materials and Methods

Approximately 20 mg of each salt was weighed into 3×2 mL push cap vials.0.1 mL of buffer (Chloride pH 1.2; Acetate pH 4.5; Phosphate pH 6.8) wasadded to each vial, at 25° C. The experiment was stirred for ca. 24 h.After 2 h, the pH was measured and adjusted if required. After 24 h afinal pH measurement was taken. Any slurries were centrifuged and thesolids analysed by XRPD. The clear solutions/mother liquors wereanalysed by HPLC

Results

The results from the pH solubility study are summarized in Table 3.8:

High solubility (>96.75 mg/mL, with respect to mesembrine) was observedfor hydrochloride salt Pattern 1 and besylate Pattern 1 across all pHranges (1.2, 4.5, 6.8).

Solubility was 29.44 mg/mL for fumarate salt Pattern 1, at pH 1.2.Solids isolated from this experiment were consistent with fumaric acid.

Oiling was observed in the fumarate salt Pattern 1 experiment at pH 6.8.

TABLE 3.8 Summary of Results from pH Studies pH HPLC Target 2 h 2 h 24 hFinal Purity/ Conc./ Salt PH measured adjusted measured pH Observation %area mg/mL XRPD Hydrochloride 1.2 1.2 1.25 n/a 1.48 Clear solution97.09 >105 — Pattern 1 4.5 4.5 4.49 n/a 4.34 Clear solution 93.45 >146 —6.8 6.8 5.67 6.74 6.54 Clear solution 92.87 >122 — Besylate 1.2 1.2 1.18n/a 1.18 Clear solution 98.01 >172 — Pattern 1 4.5 4.5 5.55 4.55 3.72Clear solution 98.02 >140 — 6.8 6.8 5.53 6.75 6.80 Clear solution97.62 >97 — Fumarate 1.2 1.2 2.31 1.21 1.36 Slurry 96.31 29.44 U Pattern1 + 4 4.5 4.5 3.28 4.49 4.53 Clear solution 96.07 >24 — 6.8 6.8 3.386.71 8.01 Oil 96.78 18.98 — Key A Amorphous # Pattern number CCrystalline potential salt PC Poorly crystalline U Counterion(unreacted) — No solids

Conclusion

The besylate salt and hydrochloride salts were highly soluble at all pHranges, providing evidence that these salts would be suitable fordevelopment of a pharmaceutical composition.

The fumarate salt was only able to produce a clear solution at pH 4.5.

Example 7: Stability Study Materials and Methods

A 1-week stability study was performed for hydrochloride salt Pattern 1,besylate salt Pattern 1, fumarate salt Pattern 1+4, and fumarate saltPattern 4.

Approximately 5 mg of the salt was weighed into 3×2 mL push cap vials.The 2 mL vials were then each placed inside a 20 mL vial, with the lidson or off as required. The solids were placed under the requiredstability conditions for 1 week at the following three conditions: 40°C./75% RH (open); 80° C. (closed); Ambient light, temperature andhumidity (open)

After 1-week, visual observations were noted, then the solids wereanalysed by XRPD and HPLC.

Results

The results from the 1-week stability studies are summarized in Table3.9.

For hydrochloride salt Pattern 1, there was a slight increase inchemical purity observed from all stability conditions. Solids ofhydrochloride salt Pattern 1 were isolated from all stabilityconditions.

For besylate salt Pattern 1, there was no decrease in chemical purityobserved after 1 week at 40° C. /75% RH, or ambient conditions. A smalldecrease in chemical purity was observed after 1 week at 80° C. Solidsof besylate salt Pattern 1 were isolated from all conditions.

For fumarate salt Pattern 1+4, there was a slight decrease in chemicalpurity observed after 1 week at 40° C./75% RH, and the solids wereconsistent with a mixture of fumarate salt Pattern 1 and Pattern 4. Adecrease in chemical purity was observed after 1 week at 80° C., withconversion to novel fumarate salt Pattern 6. No significant decrease inchemical purity was observed after 1 week at ambient conditions, and thesolids had a XRPD diffractogram consistent with a mixture of fumaratesalt Pattern 1 and Pattern 4.

For fumarate salt Pattern 4, solids of fumarate salt Pattern 4 wererecovered from all stability conditions. There was a slight increase inchemical purity after 1-week at all stability conditions.

TABLE 3.9 Summary of Results from 1-week stability study Stability HPLC/Salt Condition Observation * % area XRPD Hydrochloride Input Faint beigesolids (BE10) 90.5 1, C Pattern 1 40° C./75% RH (open) Light brownsolids (BR8) 93.4 1, C 80° C. (closed) Light brown solids (BR8) 92.8 1,C Ambient light, temperature Faint beige solids (BE10) 91.5 1, C andhumidity Besylate Input Faint beige solids (BE10) 98.1 1, C Pattern 140° C./75% RH (open) Light brown solids (BR8) 98.0 1, C 80° C. (closed)Faint beige solids (BE10) 96.7 1, C Ambient light, temperature Faintbeige solids (BE10) 98.1 1, C and humidity Fumarate Input Faint beigesolids (BE10) 96.6 1/4, C Pattern 1 + 4 40° C./75% RH (open) Faint beigesolids (BE10 95.6 1/4, C 80° C. (closed) Light brown solids (BR8) 83.46, C Ambient light, temperature Faint beige solids (BE10 96.1 1/4, C andhumidity Fumarate Input Faint beige solids (BE10) 95.9 4, C Pattern 440° C./75% RH (open) Faint beige solids (BE10) 96.4 4, C 80° C. (closed)Faint beige solids (BE10) 96.4 4, C Ambient light, temperature Faintbeige solids (BE10) 96.4 4, C and humidity Key A Amorphous # Patternnumber C Crystalline potential salt PC Poorly crystalline U Counterion(unreacted) — No solids * From the Sigma Aldrich colour chart

Conclusion

The besylate, hydrochloride and fumarate salt Pattern 4 were stableacross all storage conditions, providing evidence that these salts wouldbe suitable for development of a pharmaceutical composition.

The fumarate salt pattern 1 and 4 produced a novel salt type on storageat 80° C. was only able to produce a clear solution at pH 4.5.

Example 8: Scale-Up of Besylate Salt Pattern 1 Materials and Methods

A scale-up of the Besylate salt Pattern 1 was undertaken in order tofully characterise the salt of this form and to undertake a polymorphscreen. An anti-solvent addition step was carried out to maximise theyield:

Approximately 4.5 g of mesembrine was weighed out and dissolved in 27.7mL of MEK, and transferred to a 100 mL vessel at 25° C. Next 1.1equivalents (2.79 g) of benzenesulfonic acid was weighed out anddissolved in 17.7 mL of MEK.

The stock solution of benzenesulfonic acid was added dropwise into thevessel, and the experiment stirred for 1 h at 25° C.

The experiment was cooled to 5° C. at 0.1° C./min. At 5° C., 15 mL ofheptane was added over 3.3 h. The final solvent system was: MEK:heptane75:25% v/v.

The experiment was then stirred at 5° C. for ca. 12 h. Crusting wasobserved on the walls of the vessel, and this was manually re-introducedinto the slurry.

The solids were isolated by Buchner filtration and washed with 5 mL ofMEK:heptane 75:25% v/v.

Prior to isolation, a sub-sample of solids was taken by centrifugationfor XRPD analysis. The solids were dried at 40° C. under vacuum for ca.29 h and used for further analysis and polymorph screening.

The besylate salt was then lyophilised to prepare amorphous materialusing the following procedure.

Approximately 50 mg of besylate salt Pattern 1 was weighed into a 2 mLpush cap vial. The solids were dissolved in 0.5 mL of water, and thenfrozen at −20° C. Once frozen, the material was lyophilized. Theisolated solids were then analysed by XRPD, TGA/DSC and HPLC.

Results

An XRPD of the solids from the scale-up experiment was prepared asdetailed in FIG. 18 . Details of the peaks are found in Table 3.10below.

TABLE 3.10 XRPD Peaks for Besylate Salt Pattern 1 Pos. Height FWHMd-spacing Rel. [° 2θ ] [cts] Left [° 2θ] [Å] Int. [%] 9.2289 699.400.0895 9.58280 43.47 11.0523 643.46 0.0512 8.00559 39.99 11.1405 761.350.0768 7.94241 47.32 12.2729 481.42 0.0895 7.21196 29.92 12.7303 1215.720.1023 6.95387 75.56 13.4672 684.26 0.1151 6.57500 42.53 15.4051 286.370.1023 5.75196 17.80 16.6247 950.78 0.1023 5.33263 59.09 18.5019 121.960.1023 4.79562 7.58 19.5198 651.77 0.1151 4.54778 40.51 19.7897 242.770.0768 4.48634 15.09 20.2578 240.37 0.1023 4.38373 14.94 20.6618 756.890.1023 4.29892 47.04 21.1937 670.33 0.1407 4.19221 41.66 21.5953 542.410.1023 4.11516 33.71 22.3994 126.07 0.1535 3.96921 7.84 22.8769 212.170.0768 3.88744 13.19 23.2193 390.41 0.0640 3.83088 24.26 23.7753 881.900.1279 3.74253 54.81 24.1351 372.81 0.0768 3.68755 23.17 24.6446 1608.970.1279 3.61245 100.00 25.6285 243.78 0.1151 3.47596 15.15 26.1758 202.330.1279 3.40451 12.58 27.9112 140.83 0.1279 3.19666 8.75 28.3172 287.920.0895 3.15174 17.89 28.6091 117.85 0.1535 3.12024 7.32 29.2966 84.220.1791 3.04857 5.23 31.1585 113.96 0.1535 2.87052 7.08 32.4190 24.170.2047 2.76173 1.50 32.9906 98.16 0.1023 2.71517 6.10 33.8751 115.380.1279 2.64627 7.17

The solids were isolated with an 80% yield. The isolated solids had achemical purity of 96.4% area, by HPLC.

The theoretical yield, based on losses to mother liquor and washliquors, was 99%. The mother liquor and wash liquor concentrations were0.3 mg/mL.

FIG. 19 demonstrates the lyophilized material was predominantlyamorphous, there was one peak at 24.7°2θ in the isolated material whichis consistent with besylate salt Pattern 1.

Conclusion

The mesembrine besylate salt Pattern 1 was fully characterised by theXRPD analysis demonstrating a novel form of mesembrine salt withsuperior properties which are able to be lyophilised to produce anamorphous material suitable for use in the preparation of pharmaceuticalcompositions.

Example 9: Solubility Screen for Mesembrine Besylate Salt Materials andMethods

The amorphous besylate salt prepared in Example 8 was used in asolubility screen as follows:

Known volume aliquots of solvent system were added to each vial, withheating at 40° C. between each addition for ca. 5 minutes. Addition ofsolvents was continued until either dissolution was observed, or until100 volumes (ca. 1 mL) had been added.

After the solvent addition was complete, the experiments were stirred at40° C. for ca. 16 h. Slurries were stirred at 40° C. and clear solutionswere left to evaporate at 40° C., at ambient pressure for ca. 24 h. Thesolids were isolated by centrifuge filtration and were analysed by XRPD.

Clear solutions were left to evaporate at 40° C. under vacuum, for ca.72 h. Any solids isolated were analysed by XRPD.

Results

The results of the solubility screen are summarized in Table 3.11 below.

TABLE 3.11 Solubility screen of amorphous besylate salt No. SolventSystem/% v/v Solubility/mg/mL XRPD 1 1,4-Dioxane <10 A 2 1-Butanol <10 A3 1-Propanol ca.31 1, C 4 2-Ethoxyethanol ca.44 1, PC 5 2-Methyl THF <101, C 6 2-Propanol <10 1, C 7 2-Propanol:Water (50:50) >500 n/a 82-Propanol:Water (75:25) ca.167 n/a 9 Acetone <10 1, C 10 Acetone:Water(90:10) >500 1, PC 11 Acetonitrile >500 A 12 Anisole <10 A 13 ButylAcetate <10 A 14 Dichloromethane (DCM) ca.250 n/a 15 Diisopropyl ether<10 n/a 16 Dimethylsulfoxide (DMSO) >500 n/a 17 Ethanol ca.44 1, C 18Ethanol:Water (50:50) >500 n/a 19 Ethanol:Water (90:10) >500 1, PC 20Ethyl Acetate <10 1, C 21 Heptane <10 n/a 22 Isopropyl Acetate <10 1, PC23 Methanol >500 n/a 24 Methylethyl Ketone (MEK) <10 1, PC 25Methylisobutyl Ketone (MiBK) <10 A 26 N,N-Dimethylacetamide (DMA) >5002, C 27 N,N-Dimethylformamide (DMF) ca.250 1, C 28 N-Methylpyrrolidone(NMP) >500 n/a 29 tert-Butylmethyl Ether (tBME) <10 n/a 30Tetrahydrofuran (THF) <10 A 31 Toluene <10 A 32 Water >500 n/a Key AAmorphous 1 Besylate salt Pattern 1 C Crystalline salt 2 Besylate saltPattern 2 PC Poorly crystalline

The solubility screen demonstrated that in addition to the besylate saltpattern 1 an additional salt pattern 2 was formed in DMA.

Conclusion

The amorphous material is soluble in many solvent systems and is capableof forming an additional polymorph described as besylate salt pattern 2.

Example 10: Polymorph Screen for Mesembrine Besylate Salt Materials andMethods

The amorphous besylate salt prepared in Example 8 was used as the inputfor this set of experiments.

Approximately 1.25 g of besylate salt was weighed and dissolved in 17.5mL of water. The solution was split across 25×2 mL vials and frozen. Thefrozen material was lyophilized for ca. 24 h. The isolated solids weredried at 20° C. under vacuum for ca. 24 h. The dried solids wereanalysed by XRPD.

Solvent was added to each vial to obtain a slurry. If dissolution wasobserved more amorphous material was added to try and produce a slurry.

The experiments were temperature cycled between 30° C. and 5° C. at 0.1°C./min with 1 h holds at 30° C. and 5° C., for 48 h. After 48 h, solidswere isolated from slurries at 30° C. by centrifuge filtration andanalysed by XRPD. The solids were then dried at 40° C. under vacuum for18 h. The clear solutions were cooled to 5° C. over 10 minutes and heldfor 30 minutes to attempt to induce precipitation.

Results

The results of the solubility screen are summarized in Table 3.12 below.

TABLE 3.12 Polymorph screen of amorphous besylate salt Observation atXRPD No. Solvent System/% v/v end of experiment (damp) 1 1,4-DioxaneSlurry 1, C 2 1-Butanol Clear solution — 3 1-Propanol Slurry 2, C 42-Ethoxyethanol Slurry 2, C 5 2-Methyl THF Slurry 1 + 2, C 6 2-PropanolSlurry 2, C 7 2-Propanol:Water (75:25) Clear solution — 8 Acetone Slurry1 + 2, C 9 Acetonitrile:Ethyl acetate (50:50) Slurry 2, C 10 AnisoleSlurry 1 + 2, C 11 Dichloromethane (DCM) Slurry 2, C 12 DCM:tBME (50:50)Slurry 2, C 13 DMSO:Ethyl acetate (50:40) Clear solution — 14 EthanolClear solution — 15 Ethanol:Water (95:5) Slurry 2, C 16 Ethyl acetateSlurry 1, C 17 Isopropyl Acetate Slurry 1 + 2, C 18 Methanol:Ethylacetate (50:50) Clear solution — 19 Methylethyl Ketone (MEK) Slurry 1, C20 Methylisobutyl Ketone (MiBK) Slurry 1 + 2, C 21 NMP:tBME (50:50)Slurry 2, C 22 Tetrahydrofuran (THF) Slurry 1, C 23 Toluene Solids onvial base 2, C 24 Water Clear solution — Key — No solids 1 Besylate saltPattern 1 C Crystalline salt 2 Besylate salt Pattern 2

As shown, many of the solvent systems were able to produce either of thepolymorphs besylate salt pattern 1 or besylate salt pattern 2.Furthermore, some systems produced a mixture of both salt patterns.

The novel polymorph besylate salt pattern 2 was further characterised byXRPD as shown in FIG. 20 with peak positions and heights as specified inTable 3.13 below. A comparison of the two crystalline forms Pattern 1and Pattern 2 is additionally shown in FIG. 21 .

TABLE 3.13 XRPD Peaks for Besylate Salt Pattern 2 Pos. Height FWHMd-spacing Rel. [° 2θ] [cts] Left [° 2θ] [Å] Int. [%] 3.2456 58.55 0.614027.22259 1.10 7.4271 684.03 0.0768 11.90295 12.88 9.3019 1717.10 0.08959.50775 32.34 10.9620 2816.81 0.0895 8.07130 53.06 11.5879 1065.970.0768 7.63675 20.08 12.0804 494.59 0.0640 7.32648 9.32 12.5619 1305.280.0768 7.04670 24.59 13.4046 5309.08 0.0895 6.60552 100.00 14.5243135.29 0.1023 6.09873 2.55 15.1300 2692.72 0.1023 5.85592 50.72 15.64662424.16 0.0895 5.66374 45.66 16.0986 2570.30 0.1023 5.50572 48.4116.7575 1253.30 0.0895 5.29069 23.61 17.8837 528.70 0.0895 4.95996 9.9618.1790 2068.20 0.1023 4.88006 38.96 18.6461 2663.26 0.0895 4.7588450.16 18.8174 1606.35 0.0895 4.71591 30.26 19.5339 314.34 0.0768 4.544525.92 19.7767 438.54 0.0895 4.48928 8.26 20.7293 291.48 0.0895 4.285085.49 21.3589 2518.45 0.1151 4.16015 47.44 21.7748 573.67 0.0895 4.0816410.81 22.3767 1835.92 0.1023 3.97319 34.58 22.5713 656.22 0.0512 3.9393712.36 22.7318 843.79 0.0768 3.91192 15.89 23.2719 506.24 0.1023 3.822349.54 23.7453 4139.90 0.1151 3.74719 77.98 24.1138 719.19 0.1023 3.6907613.55 24.2892 374.86 0.0895 3.66451 7.06 24.7259 1014.59 0.0895 3.6007719.11 25.0993 2369.88 0.1023 3.54804 44.64 25.6893 456.57 0.1279 3.467878.60 26.2905 586.92 0.0624 3.38711 11.06 26.3404 538.09 0.0468 3.3892110.14 26.6066 757.13 0.1248 3.34759 14.26 26.9884 521.57 0.1092 3.301089.82 27.5335 113.88 0.1872 3.23696 2.15 28.0252 1249.17 0.1404 3.1812823.53 28.4998 96.44 0.0936 3.12937 1.82 28.9700 692.95 0.1560 3.0796413.05 29.2870 191.82 0.1248 3.04702 3.61 29.5248 138.56 0.1560 3.023022.61 30.0582 97.15 0.1092 2.97058 1.83 30.6912 98.55 0.1248 2.91073 1.8631.6949 208.95 0.1560 2.82081 3.94 32.0814 219.67 0.0780 2.78770 4.1432.5443 243.64 0.1404 2.74910 4.59 33.0909 289.14 0.0624 2.70493 5.4533.4658 66.36 0.1248 2.67548 1.25 34.4144 241.94 0.1872 2.60388 4.56

Conclusion

The amorphous material is capable of forming two different forms;characterized as besylate salt pattern 1 and besylate salt pattern 2.

Example 11: Pharmacokinetic Analysis of Mesembrine Besylate SaltMaterials and Methods

The amorphous besylate salt prepared in Example 8 was used in additionto mesembrine free base in this example to determine the pharmacokinetic(PK) characteristics of the two.

Groups of 18 male C57BI/6J mice (weighing between 20 and 30 g) receiveda single administration of test compound (10 mg/kg; i.p.) inmethylcellulose (0.5% w/v) at a nominal concentration of 1.0 ng/mL.

Three mice from each dose group were subject to cardiac puncture undergeneral anaesthesia at 0.25, 0.50, 1.00, 1.50, 2.00 and 4.00 hrspost-dose and plasma samples generated for application of standardLC-MS/MS bioanalytical methods.

Quantification of compound concentration was derived from referencecalibration data. Pharmacokinetic data were derived from serial plasmaconcentrations.

Results

Table 4 details the PK parameters measured using either the mesembrinebesylate salt or the free base mesembrine.

FIGS. 22 and 23 detail the mean total concentrations of mesembrinebesylate salt and mesembrine free base respectively.

TABLE 4 Summary of PK parameters C_(max) T_(max) AUC_(0-inf) t_(1/2)Compound (ng/mL) (hr) (ng · hr/mL) (hr) Mesembrine 400 0.25 290 0.454free base Mesembrine 451 0.25 392 0.615 besylate salt

Both mesembrine besylate salt and free base demonstrated a short T_(max)at only 15 minutes, however the half-life of the besylate salt wassubstantially longer for the besylate salt form of mesembrine than thefree base. Here it was seen that the besylate salt had a half-life of36.9 minutes whereas the free base was 27.2 minutes, a difference ofalmost 10 minutes.

Different C_(max) and AUCs were also found for the two forms ofmesembrine. The C_(max) and AUC were both found to be larger in themesembrine besylate salt than in the free base. With respect to the AUCthe mesembrine besylate salt produced an AUC of over 100 ng.hr/mLgreater than that produced by the mesembrine free base.

Table 5 below details the blood:plasma ratio and the blood:brain ratiosobserved in both the mesembrine besylate salt and the free base in aseparate cohort of satellite animals dosed in the same way butsacrificed at t=0.25 h to obtain brain samples.

TABLE 5 Summary of mean ratios Plasma Brain Mean Mean concen- concen-blood:plasma blood:brain tration tration Compound ratio ratio (ng/ml)(ng/g) Mesembrine 0.895 0.357 317 1292 free base Mesembrine 1.023 0.395417 814 besylate salt

As detailed above, plasma concentrations of besylate salt are greaterthan those observed with free base, consistent with the observationsdescribed in Table 4. Moreover, higher brain exposure was observed inthe animals dosed with besylate salt when compared to free base. Themesembrine besylate salt resulted in a higher mean blood to plasma ratiothan the free base. Such a difference infers that the salt form is ableto enter the plasma at a greater concentration than the free base.

Conclusion

The ability of the mesembrine besylate salt to preferentially improvethe PK properties of the mesembrine free salt demonstrates theimportance of using the salt form of mesembrine in pharmaceuticalpreparations.

The improvement of PK properties demonstrated by the besylate salt formof mesembrine will enable more of the active to be absorbed and as sucha smaller dose of drug can be given. Advantages of this includes a lowercost of active ingredient due to a smaller amount being required toobtain the same effect and also potentially fewer side effects for thepatient due to a lower dose of drug being required.

Overall Conclusion

The Examples 1 to 10 presented above demonstrate that alternative saltforms of mesembrine can be formed using various counterions. The HClsalt is known in the art however there are distinct problems with thissalt form as it is only able to solubilise to form a clear solution atrelatively low concentrations of mesembrine. This physicochemicalproperty renders the hydrochloride salt unsuitable for development of apharmaceutical as only low doses of mesembrine could be delivered.

However, the besylate salt was found to be highly soluble resulting in asolubility of greater than 1000 mg/ml, providing evidence that this saltis highly suitable for development of a pharmaceutical composition.

Such a finding is surprising as often salts formed from benzenesulfonicacid are rarely found as being of use in active pharmaceuticalingredients. Hydrochloride salts are the most commonly foundpharmaceutical salts and are found in approximately 15.5% of allapproved medicinal compounds. Sodium and sulphate salts are also foundin 9% and 4% of all medicinal compounds respectively.

The finding that a besylate salt was the most soluble and stable salt incomparison to the hydrochloride and fumarate salts was additionallysurprising given the relatively high pKa of benzenesulfonic acid,particularly in comparison to hydrochloric acid. As described in Example3, a weak acid is unlikely to form a suitable salt due to the inabilityto complete the proton transfer.

In the formulation of drug compositions, it is important for the activepharmaceutical ingredient (API) to be in a form in which it can beconveniently handled and processed. This is of importance, not only fromthe point of view of obtaining a commercially viable manufacturingprocess, but also from the point of view of subsequent manufacture ofpharmaceutical formulations (e.g., oral dosage forms such as tablets)comprising the active pharmaceutical ingredient.

In the manufacture of oral drug compositions, it is important that areliable, reproducible and constant plasma concentration profile of theactive pharmaceutical ingredient is provided following administration toa patient.

Chemical stability, solid state stability, and the shelf life of theactive pharmaceutical ingredient are also very important factors in theconsideration of the form to use for preparation of the pharmaceutical.

Amorphous materials, such as mesembrine, are typically more difficult tohandle and to formulate and are often unstable. Therefore, themesembrine besylate salt provides a form which can be used in themanufacture of commercially viable and pharmaceutically acceptable drugcompositions, as it has been shown to occur as a substantiallycrystalline and stable form, which is highly soluble.

Furthermore, the data presented in Example 11 demonstrates that thebesylate salt of the invention was able to produce preferentialpharmacokinetic properties than mesembrine free base.

Numbered Embldiments

1. A mesembrine salt, wherein the salt is taken from the groupconsisting of mesembrine besylate; mesembrine phosphate; mesembrinetartrate; mesembrine fumarate and mesembrine succinate.

2. A mesembrine salt according to embodiment 1, wherein the salt ismesembrine besylate salt.

3. A mesembrine salt according to embodiment 1 or embodiment 2, whereinthe salt is in a solid form.

4. A mesembrine salt according to any of the preceding embodiments,wherein the salt is in a crystalline form.

5. A mesembrine salt according to embodiment 2, characterized by an XRPDpattern substantially similar to FIG. 18 .

6. A mesembrine salt according to embodiment 5, characterized by an XRPDpattern comprising peaks at about the positions as described in Table3.10.

7. A mesembrine salt according to embodiment 2, characterized by an XRPDpattern substantially similar to FIG. 20 .

8. A mesembrine salt according to embodiment 7, characterized by an XRPDpattern comprising peaks at about the positions as described in Table3.13.

9. A mesembrine salt according to embodiment 4, wherein the crystallineform is characterized by peaks in an XPRD pattern at 11.1±0.2, 12.7±0.2,16.6±0.2, 23.8±0.2, and 24.6±0.2°2θ.

10. The mesembrine salt according to embodiment 9, further characterizedby at least one peak selected from 9.2±0.2, 11.0±0.2, 13.5±0.2,19.5±0.2, 20.7±0.2, and 21.2±0.2°2θ.

11. The mesembrine salt according to embodiment 4, wherein thecrystalline form is characterized by peaks in a XRPD pattern at 9.2±0.2,11.0±0.2, 11.1±0.2, 12.3±0.2, 12.7±0.2, 13.5±0.2, 15.4±0.2, 16.6±0.2,18.5±0.2, 19.5±0.2, 19.8±0.2, 20.2±0.2, 20.7±0.2, 21.2±0.2, 21.6±0.2,22.4±0.2, 22.9±0.2, 23.2±0.2, 23.8±0.2, 24.1±0.2, 24.6±0.2, 25.6±0.2,26.2±0.2, 27.9±0.2, 28.3±0.2, 28.6±0.2, 29.3±0.2, 31.1±0.2, 32.4±0.2,33.0±0.2, and 33.9±0.2°2θ.

12. The mesembrine salt according to embodiment 4, wherein thecrystalline form is characterized by peaks in a XPRD pattern at11.0±0.2, 13.4±0.2, 15.1±0.2, 18.6±0.2, or 23.7±0.2°2θ.

13. The mesembrine salt according to embodiment 12, furthercharacterized by at least one peak selected from 15.6±0.2, 16.1±0.2,18.2±0.2, 21.3±0.2, or 25.1±0.2°2θ.

14. The mesembrine salt according to embodiment 4, wherein thecrystalline form is characterized by peaks in a XRPD pattern at 3.2±0.2,7.4±0.2, 9.3±0.2, 11.0±0.2, 11.6±0.2, 12.1±0.2, 12.6±0.2, 13.4±0.2,14.5±0.2, 15.1 ±0.2, 15.6±0.2, 16.1±0.2, 16.8±0.2, 17.9±0.2, 18.2±0.2,18.6±0.2, 18.8±0.2, 19.5±0.2, 19.8±0.2, 20.7±0.2, 21.3±0.2, 21.8±0.2,22.4±0.2, 22.6±0.2, 22.7±0.2, 23.3±0.2, 23.7±0.2, 24.1±0.2, 24.3±0.2,24.7±0.2, 25.1±0.2, 25.7±0.2, 26.3±0.2, 26.6±0.2, 27.0±0.2, 27.5±0.2,28.0±0.2, 28.5±0.2, 29.0±0.2, 29.3±0.2, 29.5±0.2, 30.0±0.2, 30.7±0.2,31.7±0.2, 32.1±0.2, 32.5±0.2, 33.1±0.2, 33.5±0.2, and 34.4±0.2°2θ.

15. A process for the preparation of a mesembrine salt comprising thesteps of:

-   -   a) Dissolving mesembrine in a solvent;    -   b) Addition of the appropriate counterion to the mesembrine        solution under temperature cycling conditions; and    -   c) Isolation of solids comprising the mesembrine salt.

16. A process according to claim 15, wherein the counterion of step b)is benzenesulfonic acid.

17. A pharmaceutical preparation comprising a mesembrine salt, whereinthe salt is taken from the group consisting of mesembrine besylate;mesembrine phosphate; mesembrine tartrate; mesembrine fumarate andmesembrine succinate.

18. A pharmaceutical preparation according to embodiment 17, wherein thesalt is mesembrine besylate.

19. A pharmaceutical preparation according to either embodiment 17 orclaim 18, wherein the preparation produces an elevated blood level ofmesembrine of between 80% and 125% compared to those obtained with apharmaceutical preparation not comprising a salt form of mesembrine.

20. A mesembrine salt for use in the treatment of a disease, wherein thesalt is taken from the group consisting of mesembrine besylate;mesembrine phosphate; mesembrine tartrate; mesembrine fumarate andmesembrine succinate.

21. A mesembrine salt for use according to embodiment 20, wherein thesalt is mesembrine besylate.

1. A mesembrine salt, wherein the salt is mesembrine besylate;mesembrine phosphate; mesembrine tartrate; mesembrine fumarate andmesembrine succinate.
 2. The mesembrine salt according to claim 1,wherein the salt is mesembrine besylate salt.
 3. The mesembrine saltaccording to claim 1, wherein the salt is in a solid form.
 4. Themesembrine salt according to claim 1, wherein the salt is in acrystalline form.
 5. The mesembrine salt according to claim 2,characterized by an XRPD pattern of FIG. 18 .
 6. The mesembrine saltaccording to claim 5, characterized by an XRPD pattern comprising peaksat about the positions as described in Table 3.10.
 7. The mesembrinesalt according to claim 2, characterized by an XRPD pattern of FIG. 20 .8. The mesembrine salt according to claim 7, characterized by an XRPDpattern comprising peaks at about the positions as described in Table3.13.
 9. A crystalline form of mesembrine characterized by peaks in anXPRD pattern at 11.1±0.2, 12.7±0.2, 16.6±0.2, 23.8±0.2, and 24.6±0.2°2θ.10. The crystalline form of mesembrine according to claim 9, furthercharacterized by at least one peak selected from 9.2±0.2, 11.0±0.2,13.5±0.2, 19.5±0.2, 20.7±0.2, and 21.2±0.2°2θ.
 11. The crystalline formof mesembrine according to claim 9, wherein the crystalline form ischaracterized by peaks in a XRPD pattern at 9.2±0.2, 11.0±0.2, 11.1±0.2,12.3±0.2, 12.7±0.2, 13.5±0.2, 15.4±0.2, 16.6±0.2, 18.5±0.2, 19.5±0.2,19.8±0.2, 20.2±0.2, 20.7±0.2, 21.2±0.2, 21.6±0.2, 22.4±0.2, 22.9±0.2,23.2±0.2, 23.8±0.2, 24.1±0.2, 24.6±0.2, 25.6±0.2, 26.2±0.2, 27.9±0.2,28.3±0.2, 28.6±0.2, 29.3±0.2, 31.1±0.2, 32.4±0.2, 33.0±0.2, and33.9±0.2°2θ.
 12. A crystalline form of mesembrine characterized by peaksin an XPRD pattern at 11.0±0.2, 13.4±0.2, 15.1±0.2, 18.6±0.2, or23.7±0.2°2θ.
 13. The crystalline form of mesembrine according to claim12, further characterized by at least one peak selected from 15.6±0.2,16.1±0.2, 18.2±0.2, 21.3±0.2, or 25.1±0.2°2θ.
 14. The crystalline formof mesembrine according to claim 12, wherein the crystalline form ischaracterized by peaks in a XRPD pattern at 3.2±0.2, 7.4±0.2, 9.3±0.2,11.0±0.2, 11.6±0.2, 12.1±0.2, 12.6±0.2, 13.4±0.2, 14.5±0.2, 15.1 ±0.2,15.6±0.2, 16.1±0.2, 16.8±0.2, 17.9±0.2, 18.2±0.2, 18.6±0.2, 18.8±0.2,19.5±0.2, 19.8±0.2, 20.7±0.2, 21.3±0.2, 21.8±0.2, 22.4±0.2, 22.6±0.2,22.7±0.2, 23.3±0.2, 23.7±0.2, 24.1±0.2, 24.3±0.2, 24.7±0.2, 25.1±0.2,25.7±0.2, 26.3±0.2, 26.6±0.2, 27.0±0.2, 27.5±0.2, 28.0±0.2, 28.5±0.2,29.0±0.2, 29.3±0.2, 29.5±0.2, 30.0±0.2, 30.7±0.2, 31.7±0.2, 32.1±0.2,32.5±0.2, 33.1±0.2, 33.5±0.2, and 34.4±0.2°2θ.
 15. A pharmaceuticalcomprising the mesembrine salt according to claim
 1. 16. Apharmaceutical comprising the mesembrine salt according to claim
 9. 17.A pharmaceutical comprising the mesembrine salt according to claim 12.